Дисертації з теми "DNA thermodynamics"
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1
Lee, Se Il. "Statistical thermodynamics of virus assembly." Diss., Georgia Institute of Technology, 2010. http://hdl.handle.net/1853/33900.
Повний текст джерелаАнотація:
Experiments show that MgSO4 salt has a non-monotonic effect as a function of MgSO4 concentration
on the ejection of DNA from bacteriophage lambda.
There is a concentration, N0, at which the minimum amount of DNA is ejected.
At lower or higher concentrations, more DNA is ejected. We propose that this non-monotonic behavior
is due to the overcharging of DNA at high concentration of Mg⁺² counterions.
As the Mg⁺² concentration increases from zero, the net charge of ejected DNA changes its sign from negative to positive.
N0 corresponds to the concentration at which DNA is neutral.
Our theory fits experimental data well.
The DNA-DNA electrostatic attraction is found to be -0.004 kBT/nucleotide.
Simulations of DNA-DNA interaction of a hexagonal DNA bundle support our theory.
They also show the non-monotonic DNA-DNA interaction and reentrant behavior of DNA condensation by divalent counterions.
Three problems in understanding the capsid assembly for a retrovirus are studied:
First, the way in which the viral membrane affects the structure of in vivo assembled HIV-1 capsid is studied.
We show that conical and cylindrical capsids have similar energy at high surface tension of the viral membrane,
which leads to the various shapes of HIV-1 capsids. Secondly, the problem of RNA genome packaging inside spherical viruses
is studied using RNA condensation theory. For weak adsorption strength of capsid protein, most RNA genomes are located at the center
of the capsid. For strong adsorption strength, RNA genomes peak near the capsid surface and the amount of RNA packaged is proportional to the capsid area instead its volume. Theory fits experimental data reasonably well.
Thirdly, the condensation of RNA molecules by nucleocapsid (NC) protein is studied.
The interaction between RNA molecules and NC proteins is important for the reverse transcription of viral RNA which relates to the viral infectivity.
For strong adsorption strength of the NC protein, there is a screening effect by RNA molecules around a single NC protein.
2
Davis, Tina Marie. "Optical properties, conformation, and thermodynamics of DNA oligonucleotides." Diss., Georgia Institute of Technology, 1998. http://hdl.handle.net/1853/30389.
Повний текст джерела
3
Schubert, Frank. "Visualization, kinetics, and thermodynamics of DNA-protein interactions." [S.l.] : [s.n.], 2005. http://deposit.ddb.de/cgi-bin/dokserv?idn=976174790.
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4
Dickman, Rebekah. "Thermodynamic Effects of 5' and 3' Single Strand Dangling Ends on Short Duplex DNA." PDXScholar, 2010. https://pdxscholar.library.pdx.edu/open_access_etds/94.
Повний текст джерелаАнотація:
Differential scanning calorimetry (DSC) melting analysis was performed on 27 short double stranded DNA duplexes containing 15 to 25 base pairs and short single stranded overhangs from one to 10 bases, on both ends. Molecules have two 5' dangling ends or one 5' and one 3' dangling end. For these molecules the duplex region was incrementally reduced from 25 to 15 base pairs with increased length of the dangling ends from one to 10 bases. A third set of molecules contained 21 base pair duplexes with a four base dangling end on either the 5' or 3' end. Blunt ended duplexes from 15 to 25 base pairs were also examined and served as control duplexes. DSC melting curves were measured in solution containing 85 mM, 300 mM or 1.0 M Na+. From these measurements, thermodynamic parameters for 5' and 3' dangling ends as a function of end length were evaluated. Results showed the 5' ends were slightly stabilizing, and this stability was essentially constant with end length, while the 3' ends were generally destabilizing with increasing length of the end. This finding of lower stability for the 3' ends is consistent with results of published studies that have found 5' dangling ends to be more than or equally as stabilizing as 3' dangling ends. Our finding that 3' dangling ends are actually destabilizing for duplex DNA contrasts with published results. The 3' ends also display a stronger dependence on the [Na+]. In the lower Na+ environment the 3' ends are more destabilizing than at the higher salt environments. Analysis of the thermodynamic parameters of the dangling-ended duplexes as a function [Na+] indicated the 3' dangling end molecules behave differently compared to 5' dangling ended and blunt ended duplexes. The net counterion release per phosphate upon melting the molecules having one 5' and one 3' end was approximately 15% smaller as a function of end length compared to the duplex having two 5' ends. Further analysis of the DSC evaluated thermodynamic transition parameter, ΔHcal, and its relationship to the measured transition temperatures of the DNA molecules, provided an estimate on the excess heat capacity differences, ΔCp, between duplex and melted single strands for the dangling-ended molecules. The analysis revealed the molecules with one 5' and one 3' dangling end had very different ΔCp values compared to the blunt-ended molecule; while the molecules with two 5' ends have ΔCp that are essentially the same as the blunt-ended duplex. These observations are interpreted as differences in the interactions with Na+, solvent and the terminal base pairs of the duplex for the 5' versus 3' dangling ends.
5
Harris, Sarah Anne. "Theoretical investigations of DNA structure and dynamics." Thesis, University of Nottingham, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.368362.
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6
Fakhfakh, Kareem. "Quantifying and modeling the melting thermodynamics of chemically modified duplex DNA." Thesis, University of British Columbia, 2016. http://hdl.handle.net/2429/58855.
Повний текст джерелаАнотація:
Biological reagents that bind a target selectively and with high affinity are widely used as recognition molecules within diagnostic assays and as therapeutics, among other applications. By leveraging their Watson-Crick base pairing ability, short DNA oligonucelotides represent one class of such biological agents that is particularly well suited to analyzing specific elements of the human genome. Such analyses are routinely used by clinics to detect and manage disease, and those analyses are increasingly providing the richer data content and improved performance necessary for effective clinical decision-making by employing chemically modified nucleic acids. To date, the use of these unnatural nucleotides has largely been achieved empirically, but their growing use is motivating the development of new tools and guidelines that accelerate and improve their implementation in novel assays. This thesis describes how two experimental methods may be tailored to accurately measure the melting thermodynamics of short duplex DNA containing chemical modifications – specifically locked nucleic acids (LNAs) – and then reports on a study that used those methods to measure the thermal stabilities of a large panel of DNA duplexes containing LNA substitutions in one or both strands. Those data and insights gleaned from them are used to extend a molecular thermodynamic model, the “Single Base Thermodynamic” (SBT) model[1], to enable accurate predictions of the melting thermodynamics of short B-form DNA duplexes containing i) LNA:LNA base pair and/or ii) oppositely oriented LNA:DNA base pair structures. It is the only thermodynamic model with this ability, and its value is demonstrated through its use to guide the development of a entirely new type of quantitative real-time PCR based diagnostic assay – in this case directed against clinically relevant BRAFV600 mutations in cancer – that improves upon commercially available assays by bettering their throughput and limit of detection.Applied Science, Faculty of
Graduate
7
Verdemato, Philip Edward. "The interaction of the Ada protein with DNA : structure and thermodynamics." Thesis, University of Leicester, 2000. http://hdl.handle.net/2381/29654.
Повний текст джерелаАнотація:
The C-terminal domain of the E. coli Ada protein (Ada-C) aids the maintenance of genomic integrity by efficiently repairing pre-mutagenic O6-alkylguanine lesions in DNA. The aim of this thesis was to discern the manner in which Ada-C binds and repairs DNA. The research was pursued from both structural and thermodynamic perspectives, to obtain a model of the DNA-binding process. The production of recombinant Ada-C from E. coli culture was enhanced through a combination of rational media design, E. coli strain choice, and the employment of a growth strategy for maximising cell density prior to induction of protein expression. Nuclear Magnetic Resonance (NMR) studies mapped the DNA-binding site to the recognition helix of the helix-turn-helix motif and a loop region (residues 149-155) known as the 'wing'. Using this binding interface, and in the absence of a large conformational change in the protein upon DNA-binding, it was found that an O6 meG lesion was inaccessible to active site nucleophile Cys 144 when the lesion remained stacked within the DNA duplex. This lesion could enter the active site by being rendered extrahelical, or "flipping". The DNA-binding process was shown to be entropically driven, whilst the demethylation reaction provoked an exothermic heat change. At millimolar concentrations, methylation of Ada-C led to a loss of structural integrity. Duplex DNA containing an O6meG lesion had a lower enthalpy of melting than unmethylated DNA. This, along with 19F NMR work on modified DNA, led to the postulation that O6meG might induce localised duplex melting. This would certainly aid the base-flipping process.
8
Singleton, Scott F. Dervan Peter B. Dervan Peter B. "The thermodynamics of oligonucleotide-directed triple helix formation at single DNA sites /." Diss., Pasadena, Calif. : California Institute of Technology, 1995. http://resolver.caltech.edu/CaltechETD:etd-10242007-090557.
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Kilpatrick, Nancy A. "Binding of Bisbenzamidines with AT Rich DNA: A Thermodynamic Study." Digital Archive @ GSU, 2011. http://digitalarchive.gsu.edu/chemistry_theses/41.
Повний текст джерелаАнотація:
Diamidines are small molecules that generally possess antiparasitic properties and bind preferentially to the minor groove of AT rich DNA. With the goal of getting a better understanding of the thermodynamic driving forces and binding affinities, a series of pentamidine analogs were investigated with various AT rich DNA by ITC, UV-Vis and fluorescence spectroscopic methods. Findings suggest that the substitution of the linker oxygen of pentamidine to a nitrogen slightly improves the binding affinity. All of the investigated compounds are entropically driven at 25 oC with non-alternating AT DNA. Additionally, the increased fluorescence of the nitrogen and sulfur linked analogs will enable future work to be done with fluorescence microscopy to help determine if and where these compounds accumulate in the target organism.
10
Davis, Michael L. "Excimer-Monomer Switching Molecular Beacon: The Study on Synthetic Cryptosporidum DNA Detection, Thermodynamics, and Magnesium Effects." DigitalCommons@USU, 2014. https://digitalcommons.usu.edu/etd/2191.
Повний текст джерелаАнотація:
Cryptosporidium parvum is a deadly waterborne protozoan parasite that invades the gastrointestinal tract of humans and causes severe to life-threatening gastro enteric disease. Due to the ubiquitous nature of Cryptosporidium parvum in the world’s water, it is necessary to determine the source of an outbreak. Rapid detection and identification of various genotypes of Cryptosporidium are a valuable goal in determining the source of the pathogen in a human epidemic. Exploitation of gene sequences specific to species is a powerful tool detecting pathogens.
Molecular beacons are one of these tools for high selectivity and specificity detection of DNA and RNA. Molecular beacon is a single strand of DNA that forms a stem and loop structure, where the stem holds the DNA together and the loop detects a target sequence. This molecular beacon detection is determined by the changes of fluorescent emissions of fluorescent dye linked to the ends of the stem. In this thesis work, a new and novel molecular beacon was designed to detect the specific sequences from the heat shock protein gene of Cryptosporidium parvum that infects humans. This probe is synthesized by the conjugation of pyrene molecules to both ends of the stem which leads to a unique feature of pyrene excimer-monomer switching molecular beacon upon the hybridization of the loop sequence with the target DNA sequence.
This thesis systematically investigates the physical binding (e.g., quantum yield) and thermodynamic properties, including enthalpy, entropy, and free energy of this excimer-monomer switching molecular beacon in the presences of complimentary, mismatched, and damaged DNA, respectively, in the three phases: phase one is the molecular beacon in the stem and loop structure, phase two is the molecular beacon hybridized to its target DNA, and phase three is the molecular beacon in a random coil. The effect of magnesium concentration on the binding and thermodynamic properties was also investigated. Finally, as a comparison, a conventional fluorescence resonance energy transfer-based molecular beacon with a fluorophore at the 5’ end and quencher at the 3’ end was used to assess selectivity and sensitivity in detection of DNA-DNA hybridization.
11
Butcher, David S. "Thermodynamics and Kinetics of Ligand Photodissociation in Heme Proteins and Formation of DNA i-Motif." FIU Digital Commons, 2017. http://digitalcommons.fiu.edu/etd/3259.
Повний текст джерелаАнотація:
Heme proteins carry out a diverse array of functions in vivo while maintaining a well-conserved 3-over-3 α-helical structure. Human hemoglobin (Hb) is well-known for its oxygen transport function. Type 1 non-symbiotic hemoglobins (nsHb1) in plants and bacterial flavohemoglobins (fHb) from a variety of bacterial species have been predicted to carry out a nitric oxide dioxygenase function. In nsHb1 and fHb this function has been linked to protection from nitrosative stress. Herein, I combine photoacoustic calorimetry (PAC), transient absorption spectroscopy (TA), and classical molecular dynamics (cMD) simulations to characterize molecular mechanism of diatomic ligand interactions with a hexa-coordinate globin from plant (rice hemoglobin), bacterial flavohemoglobins and human hemoglobin.
In rice type 1 non-symbiotic hemoglobin (rHb1), the dynamics and energetics of structural changes associated with ligand photodissociation is strongly impacted by solvent and temperature, namely CO escape from the protein matrix is slower at pH = 6.0 compare to neutral pH (ns) due to the CD loop reorganization which forms a pathway for ligand escape. In human hemoglobin, exogenous allosteric effectors modulate energetics of conformational changes associated with the CO and O2 escape although the effectors impact on rate constants for ligand association is small. The conformational dynamics associated with ligand photorelease from fHbs from Cupriavidus necator (FHP) and Staphylococcus aureus (HMPSa) are strongly modulated by the presence of azole drugs indicating that drug association modulates structural properties of the heme binding pocket.
In addition, we carried out a study of the formation of the DNA intercalated motif (i-motif). The formation of the structure is strongly favored at acidic pH; therefore, PAC was combined with a 2-nitrobenzaldehyde pH-jump to probe formation of the i-motif on fast timescales. i-Motif folding is two-step process with the initial protonation of cytosine residues being endothermic with ΔHfast=8.5 ± 7.0 kcal mol-1 and ΔVfast=10.4 ± 1.6 mL mol-1 and subsequent nucleation/i-motif folding (τ = 140 ns) with ΔHslow=-51.5 ± 4.8 kcal mol-1 and ΔVslow=-6.6 ± 0.9 mL mol-1. The above results indicate that PAC can be employed to study diverse biochemical reactions such as DNA folding, drug binding and ligand photorelease from proteins.
12
Streu, Kristina. "Structure, Thermodynamics, and Dynamical Properties of Nucleic Acids, Proteins, and Glass-Forming Liquids." Thesis, Boston College, 2016. http://hdl.handle.net/2345/bc-ir:107098.
Повний текст джерелаАнотація:
Thesis advisor: Udayan MohantyThe stabilization of particular conformations of protein and nucleic acid structure is believed to play an important role in many important biological functions. In chapter one, the α -helical conformation and structural stability of single and double stapled all- hydrocarbon cross-linked p53 peptides when bound and unbound to MDM2 are investigated. Our study provides a comprehensive rationalization of the relationship between peptide stapling strategy, the secondary structural stability, and the binding affinity of p53-MDM2 complex. In chapter two, we study counterion-mediated collapse of a strongly charged model polyelectrolyte chain by Group-II divalent metal cations using coarse-grained Brownian dynamics simulations. Polyelectrolyte effects govern the association of counterions with the chain. Large ions are less effective in counterion condensation than small ions. However, upon counterion condensation, the reduction of the backbone charge is independent of size of the metal cations. Above a threshold value of Coulomb strength parameter, counterion release entropy drives the formation of counterion-induced compact states. In chapter three, the nature of surface tension in the random first order theory of supercooled liquid is analyzed within the framework of Landau-Lifshitz fluctuation theory. We show that the surface tension of a droplet satisfies the differential equation 4πr2(dσ)+ 8πrσ(r)− Br1/2 = 0 , where B/ T = 12πkBcv , T is temperature, kB is dr Boltzmann constant, and cv is heat capacity. A consequence is that the slope of the relaxation time at the glass transition temperature, i.e., the fragility index, is expressed as the square of the ratio of heat capacity and configurational entropy of the supercooled liquid. When backbone extended nucleosides are incorporated into a double helix, a unique helical structure is formed. In chapter four, we find that the predicted stability of modified backbone DNA strands in aqueous solution is in good agreement with experimental melting temperature data. The incorporation of extended backbone nucleosides into a duplex results in elongation of the end-to-end chain distance due to the distortion of the B-DNA conformation at the mutated base-pair insertion. We also find that the modified backbone helical twist is approximately 40 degrees, larger than B-DNA helical twist and closer to the twist angle predicted for D-form DNA. The folding of RNA tertiary structure has been described as an equilibrium between partially folded I (intermediate) states, and the fully folded native conformation, or N state. RNA is highly sensitive to the ionic environment due to its negative charge, and tertiary structures tend to be strongly stabilized by Mg2+. There is a need for models capable of describing the ion atmosphere surrounding RNA with quantitative accuracy. In chapter 5, we present a generalized Manning condensation model of RNA electrostatics for studying the Mg2+-induced RNA folding of the 58mer ribosomal fragment
Thesis (PhD) — Boston College, 2016
Submitted to: Boston College. Graduate School of Arts and Sciences
Discipline: Chemistry
13
Boccongelli, Marina. "Etude expérimentale de la stabilité, sélectivité d'appariement et dynamique d'oligonucléotides DNA-DNA et LNA-DNA." Doctoral thesis, Universite Libre de Bruxelles, 2008. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/210549.
Повний текст джерелаАнотація:
Le traitement et le diagnostic de maladies d'origine génétique suscite un grand intérêt à l'heure actuelle. De par leur spécificité d'appariement avec les acides nucléiques, les oligonucléotides possèdent un grand potentiel dans ce domaine. Ils se heurtent toutefois à des limitations majeures, dont leur faible stabilité en milieu physiologique et la difficulté qu'ils ont à franchir les membranes biologiques. De nombreuses équipes de recherche s'intéressent, afin de pallier ces limitations, à la conception et à la synthèse d'oligonucléotides chimiquement modifiés. Parmi ceux-ci, les Locked Nucleic Acids (LNA), présentant une modification qui consiste en l'insertion d'un pont −O−CH2− entre l'atome C2' et l'atome C4' du sucre, constituent une famille qui semble posséder les propriétés requises. Ils sont considérés comme des candidats très prometteurs en tant qu’agents thérapeutiques et qu’outils de diagnostic du génome. La caractérisation de la stabilité et de la sélectivité d'appariement entre les LNA et les acides nucléiques naturels est, dans ce contexte, important.Dans ce travail, nous avons étudié la stabilité, la sélectivité d'appariement ainsi que la dynamique de la structure double brin d'un oligonucléotide hybride LNA-DNA, et nous avons comparé ces propriétés à celles d'un oligonucléotide DNA-DNA de même séquence. Ce dernier est constitué de 11 paires de bases formées par l'appariement du brin 5'-GCGTGTGTGCG-3' avec le brin 3'-CGCACACACGC-5'. Dans le cas de l'hybride, les nucléotides du second brin sont tous remplacés par des LNA.
La stabilité a été étudiée expérimentalement par différentes techniques :spectroscopie d'absorption UV, calorimétrie différentielle à balayage, résonance magnétique nucléaire et calorimétrie à titrage isotherme. Ces études montrent que la stabilité du duplexe hybride est plus importante que celle du naturel, et que ce phénomène s'explique par un terme entropique plus favorable pour la formation du duplexe LNA-DNA que pour la formation du duplexe DNA-DNA.
La sélectivité d'appariement a été étudiée en comparant la stabilité des deux oligonucléotides étudiés avec celle d'oligonucléotides présentant un mésappariement dans la séquence. Nos résultats montrent que la sélectivité d'appariement du brin LNA n'est pas significativement différente de celle du brin DNA. Ce résultat ne doit cependant pas être généralisé car nous n'avons testé qu'une position centrale pour le mésappariement.
L'étude de la dynamique de la structure des oligonucléotides a été effectuée par RMN et porte sur la caractérisation de la cinétique de l'ouverture individuelle des paires de bases. Nous observons que la durée de vie de l'état fermé des paires de bases G-C est supérieure dans l'oligonucléotide LNA-DNA, tandis que l'état fermé des paires A-T semble posséder une durée de vie supérieure dans l'oligonucléotide DNA-DNA.
Au cours de ce travail de thèse nous avons pu caractériser les facteurs énergétiques à la base de la stabilité accrue des oligonucléotides chimiquement modifiés de type LNA. Nous avons montré que leur sélectivité d’appariement n’est pas toujours supérieure à celle des oligonucléotides naturels et dépend des séquences impliquées. Enfin, nous avons mis en évidence les différences entre la dynamique de la structure d’un oligonucléotide possédant des LNA et celle d’un duplexe DNA.
Doctorat en Sciences de l'ingénieur
info:eu-repo/semantics/nonPublished
14
Deegan, Brian J. "Biophysical Studies of the Binding of ERα Nuclear Receptor to DNA". Scholarly Repository, 2011. http://scholarlyrepository.miami.edu/oa_dissertations/579.
Повний текст джерелаАнотація:
Estrogen receptor α (ERα) is a member of a family of ligand-modulated transcription factors that have come to be known as nuclear receptors. ERα mediates the action of estrogens and plays an integral role in a wide range of physiological processes ranging from embryonic development and morphogenesis to reproduction to cardiovascular health. Not surprisingly, malfunction of the estrogen system is associated with a host of pathological conditions such as osteoporosis, heart disease and most notably breast cancer. Essential to its functioning as a transcription factor are specific protein-DNA interactions which are mediated by the binding of the DNA-binding (DB) domain of ERα to particular DNA sequences located within target gene promoters called estrogen response elements (EREs). Here, using a diverse array of biophysical techniques, including in particular isothermal titration calorimetry coupled with molecular modeling and semi-empirical analysis, I provide new insights into the ERα-DNA interaction in thermodynamic and structural terms. My data show that the binding of the DB domain of ERα to DNA is coupled to protonation at two specific amino acids, H196 and E203. Protonation of these residues is non-trivial and is required for high affinity binding. Amino acid sequence alignment of the DB domains of the NR family suggests that this may be a hallmark feature common to the functioning of all nuclear receptors. Furthermore, I demonstrate that the DB domain can tolerate all single nucleotide substitutions within the ERE and bind in the physiologically relevant nanomolar to micromolar range. Comparative thermodynamic analysis reveals that the DB domain binds to these ERE sequences utilizing a considerable range of energetic signatures such that any one thermodynamic component of binding is not predictive of associated affinity. In addition, it is shown that nucleotide substitution results in significant changes in secondary and three-dimensional features of the oligonucleotides and may impact binding affinity. Finally, I demonstrate that the zinc-finger of the DB domain of ERα is relatively promiscuous and can accommodate several heavy-metal divalent cations. Other than zinc, only DB domains reconstituted with cobalt, cadmium and mercury were capable of binding DNA. Incorporation of the metals resulted in a wide range of CD spectroscopic features which were found not to be predictive of DNA binding capacity. Thus, isostructure does not equate to isofunction in the case of metal reconstituted DB domain of ERα. This analysis suggests that metal coordination is not likely to be required for domain folding, but rather is required to bind DNA. Taken together, this thesis provides novel insights into the physicochemical basis of a key protein-DNA interaction essential to human health and disease. My studies bear the potential to impact the development of novel therapies harboring greater efficacy coupled with lower toxicity for the treatment of disease.
15
Okonogi, Tamara Mae. "Dynamics, thermodynamics, and structural investigations of nucleic acids using site-specific spin-labeling and electron paramagnetic resonance /." Thesis, Connect to this title online; UW restricted, 2000. http://hdl.handle.net/1773/8511.
Повний текст джерела
16
Johnson, Amy. "Determining Backbone Conformations of CRE Sequence B-DNA: A Nuclear Magnetic Resonance and Mathematical Modeling Study." Scholarship @ Claremont, 2017. http://scholarship.claremont.edu/cmc_theses/1572.
Повний текст джерелаАнотація:
Nuclear Magnetic Resonance (NMR) Spectroscopy is a crucial tool for determining the structures of biological molecules. This technique can also be used to extract thermodynamic parameters of these molecules, enhancing our understanding of their biological roles. DNA is analyzed through NMR Spectroscopy in order to identify the effect of sequence on expressivity. DNA predominantly resides in BI orientation, but a second conformation, BII, also exists. DNA can switch between BI and BII backbone conformations and the likelihood of this switching is dependent upon the energetic barrier between these two sub-states. The secondary structure of DNA, and thus its adoption of BI and BII conformation, is sequence-dependent. Therefore, the identity and neighboring base pairs of a segment of DNA have a large effect on the flexibility of the backbone. Methylation also affects backbone structure. The methyl group has been shown to promote either stabilization and/or destabilization on proximate bases. This thesis uses variable temperature NMR and Mathematica modeling to determine the backbone conformations, rate of inter-conversion between BI/BII conformations, and the energetic barrier of this fluctuation for each nucleotide step in DNA dodecamers containing the CRE binding sequence. This has been a long-term goal of the Hatcher-Skeers lab, and the data from this thesis would have been added to years of flanked CRE DNA information to reveal any patterns. In this experiment, 5’-TTTC-3’ CRE DNA dodecamers underwent NMR analyses to extract backbone flexibility parameters. Additionally, the effect of methylation was studied in scans with methylated cytosine in the central CRE sequence. The TRX scale was used to predict the BII character of these sequences. Due to technical errors, the experimental results were not able to accurately represent the specific dynamics of each backbone step. However, general trends were identified, such as adherence to and veracity of the TRX scale and the effect of methylation. It was found that the %BII of the native DNA closely resembled the TRX predictions, whilst the methylated sequence did not. The largest changes in activation energy due to methylation occurred in the central CRE sequence, suggesting methylation is a localized effect. The results reflected several trends from past CRE experiments, but the data cannot be explicitly analyzed due to the technical errors.
17
Lannan, Ford. "Folding of the human telomere sequence DNA in non-aqueous and otherwise viscous solvents." Thesis, Georgia Institute of Technology, 2012. http://hdl.handle.net/1853/47598.
Повний текст джерелаАнотація:
G-quadruplex forming human telomere sequence (HTS) DNA, has been widely studied due to the telomere's implied role in biological processes, including cellular ageing and cancer physiology. The goal of these previous efforts has been to characterize the physiologically relevant structures and their stability and dynamics in order to develop therapeutic applications. Unfortunately, understanding the biologically relevant form of the human telomere DNA is complicated by the fact that HTS-derived sequences are highly polymorphic. To further complicate the issue, recent investigations have demonstrated the ability of "cell-like" co-solvents to alter the preferred G-quadruplex fold of HTS DNA. However, the origins of G-quadruplex structure selection, the relative contributions of crowding versus dehydration, and the possible effects of co-solvents on kinetically determined folding pathways remain unresolved. Towards answering these questions, I investigated HTS DNA G-quadruplex in extreme anhydrous and high viscosity conditions utilizing a deep eutectic solvent (DES) consisting of choline chloride and urea. Herein I report that the water-free DES supports an extremely stable parallel stranded structure, consistent with observations that diminished water activity is the main cause of structural transitions to the "parallel-propeller" form. Furthermore, my research shows that the highly viscous nature of the solvent enables significant diffusion based control over HTS g-quadruplex folding rates and topology, fully consistent with Kramers rate theory. To the best of my knowledge, this is the first example of the kinetic exploration of G-quadruplex folding utilizing high friction solvent; the results of which display a decreased intramolecular folding rate of HTS DNA to a never before encountered time scale on the order of days at physiological temperature. Moreover, I have demonstrated that the folding pathway of a G-quadruplex can be altered with increased solvent friction. These discoveries are important because they highlight the need to consider the viscosity when exploring the dynamics of human telomeres specifically drug binding and folding of G-quadruplexes in vivo where cellular viscosity has been reported to be as high as 140cP. Lastly, it appears that tuning solvent viscosity could prove useful to the continued study of G-quadruplex dynamics.
18
Hughesman, Curtis. "Molecular thermodynamics of the stability of natural, sugar and base-modified DNA duplexes and its application to the design of probes and primers for sensitive detection of somatic point mutations." Thesis, University of British Columbia, 2012. http://hdl.handle.net/2429/43727.
Повний текст джерелаАнотація:
Cancer is characterized as a genetic disease associated with acquired somatic
mutations, a majority of which consist of only a single base change and are commonly
referred to as somatic point mutations (SPM). Real-time quantitative polymerase-chain
reaction (qPCR) techniques using allele specific (AS) probes or primers are widely used in
genotyping assays to detect commonly known single nucleotide polymorphisms (SNP), and
also have the potential to detect SPMs, provided the required analytical sensitivity and
specificity can be realized. One strategy to establish the necessary performance is to
introduce nucleotide analogs such as Locked Nucleic Acids (LNAs) into AS probes or
primers; however the successful design requires a fundamental understanding of both the
thermodynamics and kinetics of LNA-DNA heteroduplexes. Melting thermodynamic studies
of DNA duplexes and LNA-DNA heteroduplexes were therefore carried out using both
ultraviolet (UV) spectroscopy and differential scanning calorimetry (DSC) to quantify the
thermodynamics (ΔH⁰, ΔS⁰, ΔCp and Tm) associated with the helix-to-coil transition. Data
collected on DNA duplexes and DNA-LNA heteroduplexes were used to introduce
improvements in the “unified” nearest-neighbor model, and for the development of a new
model, referred to as the Single Base Thermodynamic (SBT) model that accurately predicts
the Tm for the melting of LNA-DNA heteroduplexes.
The SBT model was extended and applied to PCR conditions to design LNA-bearing
AS probes for qPCR assays to detect the clinically important SPMs KIT c.1799t>a (D816V)
and JAK2 c.1849g>t (V617F), and were found to significantly outperform standard AS
probes containing only DNA. The interaction of Taq polymerase with heteroduplexes
formed between an LNA-bearing primer and a target template were also studied and results used to generate general rules for designing LNA-bearing AS primers capable of unequivocal
detection of a rare mutant allele bearing a SPM. The method was then extended to allow
qPCR detection by Plexor™ technology and applied to create an AS primer directed against
the JAK2 V617F SPM that can detect one mutation in a background of more than 100,000
copies of the wild-type allele and which is now used by the Cancer Genetics Laboratory of
the British Columbia Cancer Agency (BCCA) to analyze patient samples.
19
Hüsler, Paul L. "Thermodynamic characterization of DNA Triple-Helical three-way junctions." Doctoral thesis, University of Cape Town, 1995. http://hdl.handle.net/11427/18282.
Повний текст джерелаАнотація:
Watson-Crick CWC) and Hoogsteen (HG) triple-helical three-way junctions were constructed from three 33-mer oligonucleotides. The same sub-set of sequences have been used in the arms of the junctions. The junctions differ primary in the arrangement of the branch point and the ends of the arms. In the case of the HG triple-helical three-way junction, the three 33-mer oligonucleotides can fold into hairpin structures, linked by a four membered cytosine loop. Each of the hairpins contain a homo-pyrimidine 10-mer single strand extension which interacts with a neighboring hairpin to form a triple-helix on lowering the pH (between 6 and 4), via Hoogsteen (HG) hydrogen bonding. Collectively this process results in the formation of the branch point and the triple-helical arms. In the case of the WC triple-helical three-way junction, the three 33-mer oligonucleotides interact with a neighboring oligonucleotide to form a duplex. Collectively this process leads to the formation of a double-helical three-way junction with each arm containing a 9-mer homo-pyrimidine extension connected by four cytosines. Each of the single strand extensions can mutually fold back onto the duplex arms converting each arm into a triplex. The WC and HG triple-helical three-way junctions were characterized by gel electrophoresis, temperature gradient gel electrophoresis, circular dichroism (CD), uv melting, and differential scanning calorimetry (DSC). In both structures; arm A contained exclusively TAT triad bases, while arms B and C contained an increasing number of CGC+ triads, respectively. To counteract possible crowding at the branch point the 5' sequence was shortened by one base. The assembly of the completely folded structure was found to be spontaneous if an appropriate ionic strength and pH range was chosen. A separate set of isolated arms has been investigated to elucidate the role each arm plays in the complete structure. Comparing the summed-up properties of these arms with the data obtained for the integral three-way junctions, it is obvious that the we three-way junction is partly distorted at the branch point, in line with observation obtained from double-helical three-way junctions, while the HG threeway junction is completely ordered. A set of mathematical models has been developed to describe the thermal unfolding of the multi-strand DNA structure and to identify the intermediate states. Presented is a formalism, starting from the grand partition function, that describes the effects of pH on the thermal stability of triple-helices. The formalism can be used over a wide pH range. It covers nearest neighbor electrostatic effects of closely spaced cytosines in the Hoogsteen and Watson & Crick strands. A procedure is employed to predict enthalpy and entropy changes for triplex formation. The obtained values are in good agreement with the results obtained by differential scanning calorimetry. It is the first time that multistrand, branched DNA structures of this complexity were constructed, completely described, and characterized thermodynamically.
20
Conwell, Christine C. "Kinetic and Thermodynamic Factors Govern DNA Condensate Size and Morphology." Diss., Georgia Institute of Technology, 2004. http://hdl.handle.net/1853/5213.
Повний текст джерелаАнотація:
It is well known that multivalent cations can cause DNA to condense from solution to form high-density nanometer scale particles. However, several fundamental questions concerning the phenomenon of DNA condensation remain unanswered. DNA condensation in vitro has been of interest for many years as a model of naturally occurring DNA packaging (e.g. chromatin, sperm head and virus capsid packing). More recently, DNA condensation has been of interest in optimizing artificial gene delivery, where packaging genes to an optimal size is essential to developing efficient uptake and delivery systems. The research presented in this dissertation provides an in depth biophysical study of the factors that control DNA condensate size and morphology. Millimolar changes in the ionic strength of the solution were found to alter the size of toroidal condensates. Variations in the order of addition of the counterions also significantly changed the size and morphology of the condensates. Studies were also performed to investigate the effects of static curvature and increased DNA flexibility on DNA condensation. These include the addition of static bending by sequence directed curvature, dynamic bending through protein-DNA interactions and reducing DNA persistence length by condensing single-stranded DNA. Several new models of DNA condensation are proposed based on the experimental data presented in this thesis.
21
Rumora, Amy. "Exploring DNA destabilization induced by the thymine dimer lesion using base modifying probes and thermodynamic techniques /." Connect to online version, 2007. http://ada.mtholyoke.edu/setr/websrc/pdfs/www/2007/250.pdf.
Повний текст джерела
22
Akin, Myles. "Site specific thermodynamic study of OH radical addition to DNA bases." Thesis, Georgia Institute of Technology, 2010. http://hdl.handle.net/1853/33919.
Повний текст джерелаАнотація:
In medical and health physics, we are interested in the effects of ionizing radiation on biological systems, in particular, human biology. The main process by which ionizing radiations causes damage to biological systems, is through the creation of radicals close to DNA strands. The radicals are very reactive and those created within close proximity to DNA will react with the DNA causing damage, in particular single strand or double strand breaks. This damage to the DNA can cause mutations that can kill the cell, either mitotically or apoptotically, or possibly lead to a cancerous formation. Therefore it is important to study how these radicals interact with DNA strands for a correlation between the resultant products of radical reactions and DNA strand breaks. For this study, we look at the most important radical, the OH radical and it's addition to DNA bases. We will study, through quantum chemistry, the thermodynamics of OH radical addition to the four bases, Adenine, Guanine, Cytosine and Thymine. The Jaguar program developed by Schrodinger was used for DFT calculations of the Gibbs free energy of the addition. In addition, calculations for the partial charge, HOMO's and Fukui indices were calculated and compared to experiment.
23
Malhowski, Anne M. "Examining kinetic and thermodynamic DNA destabilization caused by the cis-syn thymine dimer lesion using small molecule probes /." Connect to online version, 2005. http://ada.mtholyoke.edu/setr/websrc/pdfs/www/2005/108.pdf.
Повний текст джерела
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Bourdélat-Parks, Brooke Nicole. "Thermodynamic studies of tandem mismatches and other structural elements in Hairpin and duplex nucleic acids." Available online, Georgia Institute of Technology, 2004:, 2003. http://etd.gatech.edu/theses/available/etd-04082004-180025/unrestricted/bourdelat-parks%5Fbrooke%5Fn%5F200312%5Fphd.pdf.
Повний текст джерела
25
Mutowo, Prudence. "Thermodynamic characterization of DNA binding proteins from an extreme halophilic archaeon Haloferax volcanii." Thesis, University of Nottingham, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.478927.
Повний текст джерела
26
Fix, Lydia. "Structural and thermodynamic investigations of membrane associated polypeptides and peptide/DNA transfection complexes." Université Louis Pasteur (Strasbourg) (1971-2008), 2006. https://publication-theses.unistra.fr/public/theses_doctorat/2006/FIX_Lydia_2006.pdf.
Повний текст джерелаАнотація:
Les peptides constituent un moyen simplifié d’étudier le fonctionnement des protéines et des moyens thérapeutiques potentiellement très intéressants. Ce travail a permis l’investigation de deux peptides, le peptide synthétique LAH4 et le domaine T de la protéine diphtérique en interaction avec d’autres macromolécules. Le peptide LAH4 conçu et synthétisé par notre laboratoire, présente les propriétés générales des peptides amphipathiques et se lie également fortement à l’ADN permetant son transfert dans les cellules en tant que vecteur de transfection. Dans le but de mieux comprendre l’activité du peptide LAH4 pendant la transfection, j’ai examiné ses propriétés biophysiques par ITC (isothermal titration calorimetry), CD (Circular Dichroism) et RMN du solide. Les resultats montrent que la structure en hélice- du peptide est maintenue après complexation de l’ADN. A pH neutre des liaisons électrostatiques lient les molécules de façon non spécifique et un ratio très élevé de LAH4 est requis pour la saturation et la condensation de l’ADN. A plus faible pH des interactions électrostatiques et des contributions hydrophobes stabilisent le complexe peptide/ADN, et le ratio de saturation est réduit de presque moitié. Les données concordent à l’élaboration d’un model d’action du peptide LAH4 pendant les premières étapes de la transfection. En parallèle, nous avons developé une strategie d’expression du LAH4 chez E. Coli afin de le marquer 13C uniformément pour son étude par RMN. Des études par RMN du solide ont été entreprises sur le domaine T de la toxine diphtérique afin de comprendre la topologie dans des vésicules membranaires. Des échantillons de domaine T uniformément marqués en 15N ont été préparés en reconstituant les conditions présentes dans l’endosome de façon très simplifiée. Les résultats montrent un rôle important joué autant par le lipide anionique POPG que par le pH dans l’insertion du peptide dans la membrane et dans l’interruption de celle-ciMembrane associated proteins and peptides constitute a privileged medical target. Some of them also present also an important potential in therapeutics. This work has permitted the investigation of two peptides, the synthetic peptide LAH4 and the diphtherias toxin T domain in interaction with other macromolecules. The LAH4 peptide designed and synthesized in our laboratory, presents all the general properties of amphipathic peptides and also binds strongly to the DNA allowing its transfer into the cells. In order to better understand the activity of the LAH4 peptide during transfection, I have examined its biophysical properties by ITC (Isothermal Titration Calorimetry), CD (Circular Dichroism) and solid-state NMR. The resultats show that the -helical structure of the peptide is maintained after DNA complexation. At neutral pH, the molecules are bound in an electrostatic non-specific manner and a high ratio of LAH4 is required for DNA saturation and condensation. At low pH electrostatic interactions and hydrophobic contributions stabilize the complex and the saturation ratio is reduced. The data lead to the elaboration of a model of action for the LAH4 peptide during the first steps of transfection. In parallel, we have developed a strategy of expression of the LAH4 peptide in E. Coli in order to uniformly label the peptide 13C for its study by NMR. Solid state NMR studies have been undertaken on the diphtheria toxin domain T in order to investigate its topology inside membrane vesicles. The samples of 15N uniformly labeled T domain were prepared in a simplified system mimicking endosomal conditions. The data show an important role of the pH and of the anionic lipid POPG in the peptides membrane insertion and interruption
27
Lorenzo, Ros Sara de. "Single molecule studies in a temperature-jump optical trap." Doctoral thesis, Universitat de Barcelona, 2015. http://hdl.handle.net/10803/286174.
Повний текст джерелаАнотація:
In the field of biophysics, the study of the thermodynamic characteristics of biomolecules, such as DNA, RNA or proteins, allows us to understand more about the building blocks of life. The thermodynamic characterization of the biomolecule gives us clues as to their functions and capabilities inside every living organism.
The thermodynamic characterization of nucleic acids describes how temperature affects the stability and the structure of double stranded DNA. The melting temperature of DNA (T(M)) is defined as the temperature at which half of the DNA strands in a bulk solution experiment are in the double stranded DNA (dsDNA) or random coil configuration and half of the DNA are in the single-stranded DNA (ssDNA) configuration. Using T(M), it has been possible to experimentally determine the thermodynamic parameters of Delta-G, Delta-H and Delta-S. Viceversa, when the thermodynamic parameters of a given nucleic acid sequence are known, the TM can be predicted. This effect has important applications for biomolecule techniques such as PCR (Polymerase chain reaction) or sequencing.
Traditionally thermodynamic properties of DNA have been determined using bulk techniques such as calorimetry or UV absorbance. In both cases the melting temperature has been determined by changing the temperature or pH of the entire sample.
Over the past two decades single molecule force spectroscopy has been established as a powerful, accurate and bulk-complementary method of characterizing the thermodynamics of nucleic acids. Optical trapping is an experimental technique which allows force to be exerted on a micrometric particle by using the radiation pressure of light. The miniTweezers (mT) is the newest generation of optical tweezers instruments. This instrument can be used to exert and measure forces in a range between 1-200 pN and has unprecedented resolution (0.1 pN in force an around 1 nm in distance) with very high thermal and noise stability. Optical trapping is very useful in the field of molecular biology because it allows forces to be exerted on single biomolecules bonded to the micrometric particle.
This technique is used to carry out pulling experiments on single molecules allowing us to study the mechanical, thermodynamic and kinetic properties of the molecule. Mechanical melting or unzipping is a process that consists of pulling apart the two strands of the dsDNA until the base pairs are disrupted and the molecule converts into ssDNA. In this case, and in contrast to other techniques, force, rather than temperature or pH, is used to open the molecule.
Past experiments have shown that better resolution can be obtained using single molecule techniques than can be obtained using bulk experiments. Although force unzipping provides a direct estimation of Delta-G at room temperature, extracting the value of TM always requires the determination of the Delta-H and Delta-S contributions and until now has not been reliable accomplished.En el campo de la biofísica, el estudio de las características termodinámicas de las biomoléculas, como ADN, ARN o proteínas, permite conocer más sobre los componentes básicos de la vida. La caracterización termodinámica de las biomoléculas nos proporciona pistas sobre sus funciones y capacidades dentro de un organismo vivo. La caracterización termodinámica de los ácidos nucleicos describe como la temperatura afecta la estabilidad y la estructura de la doble cadena de ADN. La temperatura de melting del ADN (TM) se define como la temperatura a la cual la mitad de las moléculas de ADN disueltas en una solución se encuentran en configuración de doble cadena (dsDNA) y la otra mitad se encuentra en la configuración de cadena individual (ssDNA). Conociendo el valor de la TM es posible determinar experimentalmente los parámetros termodinámicos: Delta-G, Delta-H y Delta-S. Viceversa, cuando los parámetros termodinámicos de la secuencia de un ácido nucleico es conocido, la TM puede ser predecida. Este efecto tiene importantes aplicaciones en técnicas de biología molecular como PCR (en inglés Polymerase chain reaction) o secuenciación. Tradicionalmente las propiedades termodinámicas del ADN han sido medidas utilizando técnicas de volumen como calorimetría o absorbancia de UV. En ambos casos la TM ha sido calculada modificando la temperatura o el pH de toda la muestra. En las pasadas dos décadas, las técnicas de espectroscopía de fuerzas sobre moléculas individuales, han sido reconocidas como técnicas de un gran valor y precisión cuyos resultados en el estudio de la caracterización termodinámica pueden ser considerados perfectamente complementarios a los medidos en técnicas de volumen. La técnica de atrapamiento óptico es una técnica experimental la cual permite ejercer fuerza sobre una partícula micrométrica utilizando la presión de radiación de la luz. Las minipinzas (en inglés minitweezers) es una nueva generación a los instrumentos de pinzas ópticas. Este instrumento puede ser usado para ejercer y medir fuerzas en un rango de entre 1-200pN y con una resolución en fuerza y distancia sin precedentes. El atrapamiento óptico es muy útil en el campo de la biología molecular permitiendo ejercer fuerzas sobre biomoléculas individuales enganchadas. Esta técnica es usada para llevar a cabo experimentos de estiramiento sobre moléculas individuales permitiendo el estudio de las propiedades mecánicas, termodinámicas y cinéticas de la molécula bajo estudio. El experimento de unzipping o melting mecánico es un proceso que consiste en separar las dos hebras de la dsDNA hasta que los enlaces entre los pares de bases complementarios son deshechos y la molécula se convierte en ssDNA. En este caso la fuerza es usada como medio para abrir la molécula, en vez de la temperatura o el pH como en otras técnicas. Pasados experimentos han mostrado que podemos obtener mejor resolución utilizando técnicas de moléculas individuales que utilizando técnicas en volumen. Aunque la fuerza de unzipping nos proporciona una estimación directa de Delta-G a temperatura ambiente, para poder extraer el valor de TM requiere conocer las contribuciones de Delta-H y Delta-S y hasta ahora no ha sido posible. Para llevar a cabo una completa caracterización termodinámica de ácidos nucleicos es importante conocer ambas magnitudes (Fuerza y Temperatura). El mejor camino para hacer este análisis es llevar a cabo experimentos de unzipping sobre moléculas individuales de ADN a diferentes temperaturas. Por ello hemos desarrollado un novedoso instrumento de pinzas ópticas con un controlador de temperatura que nos permite modificar y cambiar la temperatura de manera local y rápida. Se ha usado un específico láser calentador con una longitud de onda con una alta absorción en agua que permite cubrir un amplio rango de temperaturas. Este instrumento nos permite grabar diversas curvas de fuerza/extensión para una molécula individual a varias temperaturas con una buena estabilidad térmica y mecánica. Este diseño tiene ciertas mejoras para reducir la convección, el cual ha sido un grave problema en previos equipos calentados a través de un láser. Este equipo ha sido usado para hacer experimentos de ADN, lo que nos ha permitido hacer un análisis promediado de Delta-G, Delta-S y Delta-H entre pares de bases en un rango de temperatura entre 5ºC y 50ºC.
28
Jia, Fuchao. "Thermodynamic and structural study of the interaction between Ru(bpy)2dppz 2+ and DNA." Phd thesis, Université de Strasbourg, 2013. http://tel.archives-ouvertes.fr/tel-01062684.
Повний текст джерелаАнотація:
Dans une première partie, nous mesurons l'affinité de l'interaction entre [Ru(pby)2dppz]2+ et l'ADN en utilisant la luminescence induite lors de la complexation. Nous étudions l'évolution de l'affinité lorsque la force ionique de la solution augmente. Dans une deuxième partie, nous modifions les extrémités d'un double brin d'ADN en y greffant des fluorophores. De la mesure de transfert d'énergie non-radiative entre ces fluorophores, nous étudions l'évolution de la longueur du complexe. Nous effectuons un dosage d'un double brin de 15 paires de bases d'ADN par le complexe ruthéné. Nous nous servons de la luminescence induite par l'intercalation du groupement dppz. Cependant, l'incrément de luminescence par groupement intercalé n'est pas connu, et nous ne pouvons pas le mesurer en saturant le brin d'ADN. Nous utilisons alors une technique mise au point par Nishida [Method for Measuring the Binding of Small Molecules to Proteins from Binding-Induced Alterations of Physical-Chemical Properties], dans laquelle deux titrations de deux solutions d'ADN de deux concentrations différentes sont effectuées. En utilisant le fait que, lorsque deux solutions d'ADN complexé par le composé ruthéné, possèdent la même luminescence par paire de base , le taux de complexation de ces deux solutions doit être le même, nous pouvons alors déterminer, sans hypothèse supplémentaire, le taux de complexation de l'ADN. De l'évolution de ce taux en fonction avec la concentration de ligand, nous déduisons son affinité pour l'ADN. Nous étudions maintenant le changement de longueur d'un double brin d'ADN de 15 paires de bases, modifié à ses deux extrémités par deux fluorophores : Alexa488 et Alexa568. Lorsque Alexa 488 est porté dans un état excité, il peut se désexciter en transférant de l'énergie de manière non-radiative à Alexa568, qui se désexcite alors en émettant des photons de plus faibles énergie que ceux émis par Alexa488. L'efficacité de ce transfert d'énergie peut être quantifié à partir de la mesure des intensités émises à basse et haute énergie. Elle dépend a priori de l'efficacité couplage (et en conséquence de la distance) entre les deux fluorophores. Nous effectuons des mesures de temps de vie des états excités de chacun des fluorophores. Nous avons observé que l'addition de ligand a pour conséquence une forte inhibition quenching des fluorophores. De l'analyse de l'évolution du temps de vie du fluorophore donneur d'une part et de celui du fluorophore accepteur d'autre part, nous déduisons l'évolution de l'efficacité du transfert d'énergie en fonction de la concentration de ligand. Nous confrontons les résultats obtenus par chacune de ces analyses, et en déduisons finalement, en nous servant de l'analyse de l'équilibre effectuée dans la première partie, l'évolution de la longueur de la chaîne en fonction du taux de complexation
29
Low, Lieh Yoon. "Thermodynamic studies of zinc binding and stability of nuclear hormone receptor DNA-binding domains." Thesis, University of Cambridge, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.620310.
Повний текст джерела
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Bourdelat-Parks, Brooke Nicole. "Thermodynamic studies of tandem mismatches and other structural elements in Hairpin and duplex nucleic acids." Diss., Georgia Institute of Technology, 2003. http://hdl.handle.net/1853/5387.
Повний текст джерела
31
Rossi-Gendron, Caroline. "Dynamic DNA origamis as isothermal supramolecular machines : melting dynamics, photocontrol and isothermal folding." Thesis, Sorbonne université, 2018. http://www.theses.fr/2018SORUS522.
Повний текст джерелаАнотація:
Les origamis d’ADN et les Single Stranded Tiles (SST) semblent être deux des composants les plus prometteurs du domaine des nanotechnologies d’ADN en termes de conceptions et d'applications possibles. Dans cette thèse, nous avons exploré les aspects thermodynamiques et cinétiques sous-jacents à la formation de ces objets, ainsi que de nouvelles méthodes pour construire des nanoobjets programmables dynamiques. Notamment, l'étude du processus de formation a mis en évidence la présence inutile d'ions magnésium ou de molécules tampons dans le milieu, et de nouvelles conditions de formation ont été décrites. Le processus de fusion a été caractérisé à l'aide d'une nouvelle méthode d'électrophorèse sur gel quantifiée mettant en évidence un comportement non monotone et appelant une nouvelle définition de la température de fusion des origamis. De plus, nous avons démontré que la formation et la fusion pouvaient être contrôlées par la lumière en utilisant AzoDiGua, un intercalant d’ADN photosensible mis au point précédemment par notre groupe. Cela nous a permis d'observer pour la première fois un processus d'hybridation/déshybridation contrôlé par la lumière au sein d'origamis individuels à température constante et d'obtenir ainsi un mouvement contrôlé à l'échelle nanométrique. Nous avons également mis au point une méthode originale pour la formation isotherme d’ADN origamis et de SST à température ambiante constante et en l’absence de tout agent dénaturant. Cela nous a permis d'observer pour la première fois et in situ le pliage isotherme d'origamis individuels, démontrant ainsi que l'origami peut atteindre sa forme d'équilibre final en suivant une variété de voies de pliageDNA origamis and Single Stranded Tiles (SST) appear to be two of the most promising components of the DNA nanotechnology field in terms of possible designs and applications. In this thesis, we explored the thermodynamic and kinetic aspects underlying DNA nanostructures formation as well as new practical ways to build dynamic programmable nano-objects. Notably, the study of the formation process evidenced the unnecessary presence of magnesium ions or buffering molecules in the medium, and new formation conditions have been described. The melting process triggered by temperature elevation was characterised using a new quantified gel electrophoresis method evidencing for the first time a non-monotonous behaviour and calling for a new definition of DNA origami melting temperature. Both formation and melting process were furthermore demonstrated to be controllable by light using AzoDiGua, a photosensitive DNA intercalator previously developed by our group. This allowed us to observe for the first time a light-controlled hybridisation / dehybridisation process within individual origamis at constant temperature and thus achieve a controlled motion at the nanoscale. We also established an original method for the isothermal formation of DNA origamis and SST at constant room temperature and without the presence of any denaturating agent. This allowed us to observe for the first time and in situ the isothermal folding of individual origamis, thus evidencing that origamis can reach their final equilibrium shape following a variety of folding pathways
32
Cui, Tengjiao. "Specific binding of mammalian high mobility group protein at-hook 2 to the minor groove of at-rich DNAS : thermodynamic and specificity studies." FIU Digital Commons, 2007. http://digitalcommons.fiu.edu/etd/2683.
Повний текст джерелаАнотація:
The mammalian high mobility group protein AT-hook 2 (HMGA2) is a small transcriptional factor involved in cell development and oncogenesis. It contains three "AT-hook" DNA binding domains, which specifically recognize the minor groove of AT-rich DNA sequences. It also has an acidic C-terminal motif. Previous studies showed that HMGA2 mediates all its biological effects through interactions with AT-rich DNA sequences in the promoter regions. In this dissertation, I used a variety of biochemical and biophysical methods to examine the physical properties of HMGA2 and to further investigate HMGA2's interactions with AT-rich DNA sequences. The following are three avenues perused in this study: (1) due to the asymmetrical charge distribution of HMGA2, I have developed a rapid procedure to purify HMGA2 in the milligram range. Preparation of large amounts of HMGA2 makes biophysical studies possible; (2) Since HMGA2 binds to different AT-rich sequences in the promoter regions, I used a combination of isothermal titration calorimetry (ITC) and DNA UV melting experiment to characterize interactions of HMGA2 with poly(dA-dT)2 and poly(dA)poly(dT). My iii results demonstrated that (i) each HMGA2 molecule binds to 15 AT bp; (ii) HMGA2 binds to both AT DNAs with very high affinity. However, the binding reaction of HMGA2 to poly(dA-dT)2 is enthalpy-driven and the binding reaction of HMGA2 with poly(dA)poly(dT) is entropy-driven; (iii) the binding reactions are strongly depended on salt concentrations; (3) Previous studies showed that HMGA2 may have sequence specificity. In this study, I used a PCR-based SELEX procedure to examine the DNA binding specificity of HMGA2. Two consensus sequences for HMGA2 have been identified: 5'-ATATTCGCGAWWATT-3' and 5'-ATATTGCGCAWWATT-3', where W represents A or T. These consensus sequences have a unique feature: the first five base pairs are AT-rich, the middle four to five base pairs are GC-rich, and the last five to six base pairs are AT-rich. All three segments are critical for high affinity binding. Replacing either one of the AT-rich sequences to a non-AT-rich sequence causes at least 100-fold decrease in the binding affinity. Intriguingly, if the GC-segment is substituted by an AT-rich segment, the binding affinity of HMGA2 is reduced approximately 5-fold. Identification of the consensus sequences for HMGA2 represents an important step towards finding its binding sites within the genome.
33
Marcaud, Hélène. "Etude sur les transitions coopératives et les cinétiques de transconformation associées dans les acides nucléiques." Paris 6, 1986. http://www.theses.fr/1986PA066514.
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"Thermodynamics and Kinetics of DNA Nanostructure Assembly." Doctoral diss., 2011. http://hdl.handle.net/2286/R.I.14266.
Повний текст джерелаАнотація:
abstract: ABSTRACT The unique structural features of deoxyribonucleic acid (DNA) that are of considerable biological interest also make it a valuable engineering material. Perhaps the most useful property of DNA for molecular engineering is its ability to self-assemble into predictable, double helical secondary structures. These interactions are exploited to design a variety of DNA nanostructures, which can be organized into both discrete and periodic structures. This dissertation focuses on studying the dynamic behavior of DNA nanostructure recognition processes. The thermodynamics and kinetics of nanostructure binding are evaluated, with the intention of improving our ability to understand and control their assembly. Presented here are a series of studies toward this goal. First, multi-helical DNA nanostructures were used to investigate how the valency and arrangement of the connections between DNA nanostructures affect super-structure formation. The study revealed that both the number and the relative position of connections play a significant role in the stability of the final assembly. Next, several DNA nanostructures were designed to gain insight into how small changes to the nanostructure scaffolds, intended to vary their conformational flexibility, would affect their association equilibrium. This approach yielded quantitative information about the roles of enthalpy and entropy in the affinity of polyvalent DNA nanostructure interactions, which exhibit an intriguing compensating effect. Finally, a multi-helical DNA nanostructure was used as a model `chip' for the detection of a single stranded DNA target. The results revealed that the rate constant of hybridization is strongly dominated by a rate-limiting nucleation step.Dissertation/Thesis
Ph.D. Chemistry 2011
35
"Thermodynamics and Biological Applications of DNA Nanostructures." Doctoral diss., 2014. http://hdl.handle.net/2286/R.I.25042.
Повний текст джерелаАнотація:
abstract: DNA nanotechnology is one of the most flourishing interdisciplinary research fields. Through the features of programmability and predictability, DNA nanostructures can be designed to self-assemble into a variety of periodic or aperiodic patterns of different shapes and length scales, and more importantly, they can be used as scaffolds for organizing other nanoparticles, proteins and chemical groups. By leveraging these molecules, DNA nanostructures can be used to direct the organization of complex bio-inspired materials that may serve as smart drug delivery systems and in vitro or in vivo bio-molecular computing and diagnostic devices. In this dissertation I describe a systematic study of the thermodynamic properties of complex DNA nanostructures, including 2D and 3D DNA origami, in order to understand their assembly, stability and functionality and inform future design endeavors. It is conceivable that a more thorough understanding of DNA self-assembly can be used to guide the structural design process and optimize the conditions for assembly, manipulation, and functionalization, thus benefiting both upstream design and downstream applications. As a biocompatible nanoscale motif, the successful integration, stabilization and separation of DNA nanostructures from cells/cell lysate suggests its potential to serve as a diagnostic platform at the cellular level. Here, DNA origami was used to capture and identify multiple T cell receptor mRNA species from single cells within a mixed cell population. This demonstrates the potential of DNA nanostructure as an ideal nano scale tool for biological applications.Dissertation/Thesis
Ph.D. Chemistry 2014
36
"Thermodynamics and Kinetics of DNA Tile-Based Self-Assembly." Doctoral diss., 2016. http://hdl.handle.net/2286/R.I.38474.
Повний текст джерелаАнотація:
abstract: Deoxyribonucleic acid (DNA) has emerged as an attractive building material for creating complex architectures at the nanometer scale that simultaneously affords versatility and modularity. Particularly, the programmability of DNA enables the assembly of basic building units into increasingly complex, arbitrary shapes or patterns. With the expanding complexity and functionality of DNA toolboxes, a quantitative understanding of DNA self-assembly in terms of thermodynamics and kinetics, will provide researchers with more subtle design guidelines that facilitate more precise spatial and temporal control. This dissertation focuses on studying the physicochemical properties of DNA tile-based self-assembly process by recapitulating representative scenarios and intermediate states with unique assembly pathways.
First, DNA double-helical tiles with increasing flexibility were designed to investigate the dimerization kinetics. The higher dimerization rates of more rigid tiles result from the opposing effects of higher activation energies and higher pre-exponential factors from the Arrhenius equation, where the pre-exponential factor dominates. Next, the thermodynamics and kinetics of single tile attachment to preformed “multitile” arrays were investigated to test the fundamental assumptions of tile assembly models. The results offer experimental evidences that double crossover tile attachment is determined by the electrostatic environment and the steric hindrance at the binding site. Finally, the assembly of double crossover tiles within a rhombic DNA origami frame was employed as the model system to investigate the competition between unseeded, facet and seeded nucleation. The results revealed that preference of nucleation types can be tuned by controlling the rate-limiting nucleation step.
The works presented in this dissertation will be helpful for refining the DNA tile assembly model for future designs and simulations. Moreover, The works presented here could also be helpful in understanding how individual molecules interact and more complex cooperative bindings in chemistry and biology. The future direction will focus on the characterization of tile assembly at single molecule level and the development of error-free tile assembly systems.Dissertation/Thesis
Doctoral Dissertation Chemistry 2016
37
"Thermodynamics studies of DNA: development of the next nearest-neighbor (NNN) model." 2001. http://library.cuhk.edu.hk/record=b5895899.
Повний текст джерелаАнотація:
Ip Lai Nang.Thesis (M.Phil.)--Chinese University of Hong Kong, 2001.
Includes bibliographical references (leaves 67-71).
Abstracts in English and Chinese.
ABSTRACT (ENGLISH) --- p.iii
ABSTRACT (CHINESE) --- p.iv
ACKNOWLEDGEMENTS --- p.v
TABLE OF CONTENTS --- p.vi
LIST OF TABLES --- p.viii
LIST OF FIGURES --- p.ix
LIST OF APPENDIX --- p.x
Chapter CHAPTER 1 --- INTRODUCTION --- p.1
Chapter CHAPTER 2 --- BACKGROUND --- p.3
Chapter 2.1 --- Structure of DNA --- p.3
Chapter 2.2 --- Sequence dependent stability --- p.8
Chapter 2.3 --- Thermodynamics of DNA --- p.9
Chapter 2.4 --- Model for predicting thermodynamic parameters of DNA sequence --- p.15
Chapter 2.4.1 --- The nearest-neighbor (NN) model
Chapter 2.4.1.1 --- Background --- p.15
Chapter 2.4.1.2 --- Method for predicting thermodynamic parameters --- p.16
Chapter 2.4.1.3 --- Limitation of the NN model --- p.19
Chapter CHAPTER 3 --- EXPERIMENTAL METHOD --- p.20
Chapter 3.1 --- Design of DNA sequences PAGE --- p.20
Chapter 3.2 --- DNA synthesis and purification --- p.22
Chapter 3.3 --- UV measurement --- p.23
Chapter CHAPTER 4 --- THE NEXT NEAREST-NEIGHBOR (NNN) MODEL --- p.27
Chapter 4.1 --- Method for extracting the NNN thermodynamic parameters --- p.30
Chapter 4.2 --- Discussions --- p.34
Chapter 4.2.1 --- Comparison of the NN model and the NNN model --- p.34
Chapter 4.2.2 --- The NNN effect --- p.38
Chapter 4.2.3 --- Sequence-specific local structure of DNA and the NNN effect
Chapter CHAPTER 5 --- SUMMARY AND FUTURE WORK --- p.49
APPENDIX I´ؤ XVI --- p.51
REFERENCE --- p.67
38
"Effect of 3-methylthymine on solution structures and thermodynamic stabilities of double-helical deoxyribonucleic acids." 2011. http://library.cuhk.edu.hk/record=b5894763.
Повний текст джерелаАнотація:
Zhong, Yangliu.Thesis (M.Phil.)--Chinese University of Hong Kong, 2011.
Includes bibliographical references (leaves 50-57).
Abstracts in English and Chinese.
Title Page --- p.i
Thesis Committee --- p.ii
Abstract (English Version) --- p.iv
Abstract (Chinese Version) --- p.V
Acknowledgement --- p.vi
Table of Contents --- p.viii
List of Tables --- p.X
List of Figures --- p.xii
List of Abbreviations and Symbols --- p.xiii
Chapter 1 --- Introduction --- p.1
Chapter 1.1 --- DNA methylation --- p.1
Chapter 1.2 --- Repair of m3T --- p.2
Chapter 1.3 --- Objectives of this work --- p.3
Chapter 1.4 --- DNA structure --- p.3
Chapter 1.4.1 --- Nomenclature scheme for DNA --- p.3
Chapter 1.4.2 --- Base pair scheme --- p.4
Chapter 1.4.3 --- Sugar conformation --- p.5
Chapter 1.4.4 --- Backbone conformation --- p.7
Chapter 2 --- Materials and Methods --- p.9
Chapter 2.1 --- Sample design --- p.9
Chapter 2.2 --- Sample preparation --- p.10
Chapter 2.3 --- NMR analysis --- p.10
Chapter 2.3.1 --- Resonance assignment --- p.12
Chapter 2.3.2 --- Determination of sugar conformation --- p.13
Chapter 2.3.3 --- Determination of backbone conformation --- p.14
Chapter 2.4 --- UV melting study --- p.15
Chapter 3 --- Effect of m3T on Double-Helical Structures and Stabilities --- p.17
Chapter 3.1 --- Resonance assignments --- p.17
Chapter 3.2 --- Effect of m3T on double-helical DNA structures --- p.19
Chapter 3.2.1 --- Base pairing mode --- p.19
Chapter 3.2.2 --- Sugar conformation --- p.21
Chapter 3.2.3 --- Backbone conformation --- p.22
Chapter 3.3 --- Effect of m3T on double-helical DNA stabilities --- p.25
Chapter 3.4 --- Discussion --- p.26
Chapter 3.4.1 --- Single-strand requirement in FTO repair --- p.26
Chapter 3.4.2 --- Relationship between m3T pairing structure and stability --- p.27
Chapter 4 --- Effect of m3T Mispair on Double-Helical DNA Structures and Stabilities --- p.28
Chapter 4.1 --- Resonance assignments --- p.28
Chapter 4.2 --- Effect of m3T mispair on double-helical DNA structures --- p.32
Chapter 4.2.1 --- Pairing mode of T m3T --- p.34
Chapter 4.2.2 --- Pairing mode of G m3T --- p.35
Chapter 4.2.3 --- Pairing mode of C.m3T --- p.35
Chapter 4.3 --- Effect of m3T mispair on double-helical DNA stabilities --- p.36
Chapter 4.4 --- Discussion --- p.36
Chapter 4.4.1 --- Predominant mutation --- p.37
Chapter 4.4.2 --- Relationship between m3T pairing structure and stabilities --- p.37
Chapter 5 --- Conclusion and Future Work --- p.39
Chapter Appendix I --- Proton chemical shift values (ppm) of AmT --- p.40
Chapter Appendix II --- Proton chemical shift values (ppm) of RefAT --- p.41
Chapter Appendix III --- Proton chemical shift values of NmT samples --- p.42
Chapter Appendix IV --- "Σ1' and %S of TmT, GmT and CmT" --- p.45
Chapter Appendix V --- "1H-31P HSQC spectra of (a) TmT, (b) GmT and (c) CmT" --- p.46
Chapter Appendix VI --- "1H-31P COSY spectra of (a) TmT, (b) GmT and (c) CmT" --- p.47
Chapter Appendix VII --- "31P chemical shifts, 3JH3'P and %Bi of TmT, GmT and CmT" --- p.48
Chapter Appendix VIII --- "UV melting curves of RefAT, AmT, TmT, GmT and CmT" --- p.49
References --- p.50
39
"Applications of Cooperative DNA Biosensors." Master's thesis, 2018. http://hdl.handle.net/2286/R.I.49176.
Повний текст джерелаАнотація:
abstract: Cooperativity can be used to manipulate binding affinities of DNA biosensors – improving specificity without sacrificing sensitivity; examples include tentacle probes (TPs) and cooperative primers (CPs). This thesis body of work: (1) used TPs to develop a rapid, low-cost diagnostic for detecting the point mutation leading to Navajo Neurohepatopathy (NNH) and (2) used CPs to amplify a symmetric bowtie-barcoded origami with captured t-cell receptor (TCR) α and β mRNA of a single cell.
NNH (affecting 1-in-1600 Navajo babies) is a fatal genetic disorder often caused by 149G>A mutation and is characterized by brain damage and liver disease/failure. Phoenix Children’s Hospital currently uses gene sequencing to identify the 149G>A mutation. While this process is conclusive, there are limitations, as it requires both time (3-4 weeks) and money (>$700). Ultimately, these factors create barriers that can directly impact a patient’s quality of life. Assessment of the developed TP diagnostic, using genomic DNA derived from FFPE patient liver samples, suggests nearly 100% specificity and sensitivity while reducing cost to ~$250 (including cost of labor) and providing a diagnosis within 48 hours.
TCR specificity is dependent on V(D)J recombination as well as pairing of the αβ chains. Drs. Schoettle and Blattman have developed a solution in which a bowtie-barcoded origami strand nanostructure is transfected into individual cells of a heterogeneous cell population to capture and protect αβ mRNA. When PCR of the origami template is performed with Vα, X, Vβ, and Y primers, the α and β gene segments cannot be tied back to a barcode – and paired. Assessment of the developed CPs for PCR suggests correct individual amplification using (1) Va + Xcp and (2) Vβ + Ycp primers, whereas combination of all the primers (Va, Xcp, Vb, and Ycp) suggests hybridization of the Vα + Xcp and Vβ + Ycp products due to the origami target symmetry.Dissertation/Thesis
Third Iteration for Cooperative Primers
Masters Thesis Biomedical Engineering 2018
40
Vander, Meulen Kirk A. "The thermodynamics of DNA binding and wrapping by the E. coli DNA-remodeling protein integration host factor." 2007. http://www.library.wisc.edu/databases/connect/dissertations.html.
Повний текст джерела
41
Boehm, Belinda Jayne. "A mechanistic study of the stability of triple-stranded DNA structures." Thesis, 2017. http://hdl.handle.net/2440/113110.
Повний текст джерелаАнотація:
The DNA triple helix is a non-canonical nucleic acid structure with roles in cellular regulation.
Formed when a single strand of nucleic acid, a triplex–forming oligomer (TFO),
binds in the duplex major groove, its formation is known to be involved in the onset of the
neurodegenerative disorder Friedreich’s ataxia (FRDA). Alternatively, it may have clinical
applications in the development of novel antimicrobial agents. As a means to treat FRDA,
methods to destabilise triplexes have been examined, with the binding of ligands in the minor
groove a promising starting point. On the other hand, stabilisation of these structures may
be achieved through modification of the backbone or the bases with the aim of suppressing
bacterial genes as an interesting solution to the problem of multi-drug resistance. The origins
of triplex stability are currently not well understood and a better comprehension of these
properties is therefore an important step in the development of new medicinal technologies
involving the creation or destruction of triplexes.
In examining the effect of ligand binding on triplex stability, a single minor-groove binder,
netropsin, a known triplex destabiliser, is considered. Free energy calculations on the binding
of a 15-base TFO to a duplex give good agreement with experiment and it is found that
netropsin destabilises this triplex by approximately 15 kcal/mol. This appears to be a highly
localised effect, occurring only when netropsin is bound opposite the TFO, and associated
with a decrease in the width of the minor groove. Structural distortions associated with
netropsin and TFO binding appear, therefore, to play a large role in the ligand’s ability to
destabilise triplexes.
Although netropsin is thought to bind in the minor groove, binding in the other two grooves
formed when a TFO binds, has not been examined. Here it is found that binding in both
the minor and W–H, being the larger of the two grooves formed by triplex binding, grooves
are similarly stable, with the minor groove potentially being more stable due to strong vdW
interactions.
To study the effect of TFO composition on triplex stability, the relative stabilities of purine
and pyrimidine triplexes are examined. Significant distortion in the TFO backbone is observed
in the case of the purine triplex, with Hoogsteen pairs between adenine bases failing
to form, indicating it is likely less stable. However, pyrimidine TFO binding requires significantly
more backbone rearrangement to bind to the duplex, potentially disfavouring its
formation despite it likely being the more stable triplex.
Examining the effect of changing the TFO backbone composition from DNA to RNA, for
a purine triplex, the two structures are found to have similar stability, despite reports of the
RNA purine triplex not forming experimentally. A significant change in the sugar pucker of the purine RNA backbone is found to be required for binding, potentially explaining this lack
of formation. However, no large structural differences were found between DNA and RNA
pyrimidine triplexes to explain the previously reported greater stability of the RNA structure,
suggesting that conformational change may not entirely explain the relative stabilities of the
triplexes.Thesis (M.Phil) -- University of Adelaide, School of Physical Sciences, 2018
42
Schubert, Frank [Verfasser]. "Visualization, kinetics, and thermodynamics of DNA-protein interactions / submitted by Frank Schubert." 2005. http://d-nb.info/976174790/34.
Повний текст джерела
43
Liu, An-Chen, and 劉安振. "Studies of Thermodynamics and Mechanism of DNA Hybridization by Isothermal Titration Calorimetry." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/87128189400484487764.
Повний текст джерелаАнотація:
碩士國立中央大學
化學工程與材料工程研究所
91
Abstract DNA hybridization reaction includes ssDNA structural arrangement hydrogen bond interaction, stacking interaction between base pairs, hydration step and dehydration step. The DNA hybridization reaction is mainly affected by the negatively changed phosphate group in DNA backbone. Hydrophobic interaction between the base pairs intermolecule and intramolecule of ssDNA and electrostatic interaction play important roles in DNA hybridization reaction. Specifically, before DNA hybridization, single strand DNA molecules proceeded conformational transform from unstructured single state to helical single strand state and dehydration step. Then, double helical structure was form due to hydrogen bond interaction, stacking interaction between base pairs and hydration step. Conformational transform of single strand DNA, hydrogen bond, stacking between base pairs and hydration step are exothermic and dehydration step is endothermic, basically. Therefore, DNA hybridization enthalpy is affected to different extent by the above mentioned steps. In this study, we measured the DNA hybridization enthalpy by isothermal titration calorimetry at different temperature, DNA length, GC% and salt concentration to understand DNA molecules interaction and effects of different experiment conditions and to discuss the DNA hybridization mechanism. An experimental equation of hybridization enthalpy as function of the reaction parameters was proposed in our study. Our results demonstrated that the hybridization enthalpy is exothermic. The dehydration step plays a profound role in DNA hybridization process. It also provided theoretical basis that to select hybridization conditions for application of biochip. Furthermore we indicated the difference of hybridization enthalpy in various DNA length and between perfect match and mismatch can be differentiated in a proper reaction condition. Therefore, we may possibly detect the SNP problem by isothermal titration calorimetry measurement.
44
Singleton, Scott F. "The thermodynamics of oligonucleotide-directed triple helix formation at single DNA sites." Thesis, 1995. https://thesis.library.caltech.edu/4237/1/Singleton_sf_1995.pdf.
Повний текст джерелаАнотація:
NOTE: Text or symbols not renderable in plain ASCII are indicated by [...]. Abstract is included in .pdf document.
The local triple-helical complexes formed upon binding of pyrimidine oligonucleotides to double-helical DNA offer a versatile structural motif for designing sequence specific duplex DNA-binding molecules. The experimental determination of oligonucleotide association constants as a function of solution conditions is necessary to characterize the noncovalent forces which contribute to the affinity and specificity of DNA recognition by triple helix formation. Such information will be important for the rational application of oligonucleotide-directed triple helix formation to such tasks as the manipulation of large DNAs, and the artificial modulation of biological events. We are interested in controlling the relative amount of a local triple-helical complex formed at equilibrium under conditions which approximate those found within cells. Thus, we chose to measure equilibrium constants for the formation of triple-helical complexes at single sites within larger duplex DNAs in mixed valence salt solutions near neutral pH.
Concepts borrowed from existing footprint titration methods were combined with the powerful affinity cleaving strategy in the development of the affinity cleavage titration method for the measurement of single-site DNA-binding isotherms (Chapters One and Two). The amount of site-specific cleavage of a radiolabeled plasmid DNA fragment produced by a 15mer oligonucleotide-EDTA[...]Fe is measured over an oligonucleotide concentration range covering four orders of magnitude, allowing a binding curve to be constructed. Curve-fitting a Langmuir titration isotherm to the data points affords an equilibrium binding constant, [...], that is identical within experimental uncertainty to those obtained from quantitative DNase I footprint titrations of the same oligonucleotide with and without EDTA[...]In. Further affinity cleavage titrations demonstrate that the energetic stabilization of this local triple-helical structure depends on the length of the oligonucleotide, the presence of base triplet mismatches, and the solution conditions, including the pH, the concentration and valence of cations, and the temperature.
Decreasing the length of the oligonucleotide from 15 to 11 nts reduces the stability of the corresponding triplexes by ~0.3 kcal/mol per nucleotide removed (Chapter Two). Single internal base triplet mismatches result in a destabilization of the local triple-helical structure by ~ 3 kcal/mol. The equilibrium association constants for the 15mer increased by 10-fold as the pH was decreased from pH 7.6 to pH 5.8, indicating that the corresponding triple-helical complex was stabilized by 1.4 kcal[...]mol[superscript -1] at the lower pH (Chapter Three). Equilibrium association constants for the 15mer in the presence of various concentrations of KCl,MgCl[subscript 2], and spermine tetrahydrochloride (SpmCl[subscript 4]) are reported in Chapter Four. Varying the K[...] concentration by a factor of 28 in the presence NaCl, MgCl[subscript 2], and SpmCl[subscript 4] resulted in an overall 100-fold decrease in the binding affinity from the lowest to the highest concentration. In contrast, measured binding constants increased 500-fold as the Spm[superscript 4+] concentration was increased 10-fold.
There was a modest effect on the binding constant (a 3-fold decrease) upon increasing the Mg[superscript 2+] concentration by a factor of 100. The influence of temperature on the energetics of oligonucleotide-directed triple helix formation in mixed valence salt solutions at pH 7.0 are reported in Chapter Five. In three solutions differing in their salt compositions, the equilibrium association constants decreased at least 100-fold (from >10[superscript 7]M[superscript -1] to ~ 10[superscript 5]M[superscript -1]) as the temperature was increased from 8°C to 37°C. Least squares analysis of van't Hoff plots (lnK versus 1/T) of the data revealed that in each solution the triplex is enthalpically stabilized by ~ 2 kcal per mole of base triplets.
This value for the enthalpic contribution to triplex stability, which reflects contributions from both T[...]AT and C+GC base triplets, is identical within experimental error to that measured previously using differential scanning calorimetry for the 15mer binding to 15 by within a 21-bp duplex. We collaborated with Professor Kenneth J. Breslauer of Rutgers University to make the calorimetric measurement, and the complete thermodynamic characterization of the stability and the melting behavior of the triplex are described in Chapter Six.
Taken together, the equilibrium association constants, the thermal denaturation data, and the calorimetric data reported here provide a quantitative measure of the influence of third strand length, base triplet mismatches, pH, salt, and temperature on the stability and the melting behavior of a DNA triplex. Such information should prove useful in designing triplex-forming oligonucleotides and in defining solution conditions for the effective use of triple-helical structure formation as a tool for modulating biochemical events.
45
Koh, Junseock. "Thermodynamics of interactions of Escherichia coli HU[alpha, beta] with duplex DNA." 2008. http://www.library.wisc.edu/databases/connect/dissertations.html.
Повний текст джерела
46
Holbrook, Jill Anne. "Calormetric studies of DNA helix formation and integrational host factor-DNA interactions : contribution of coupled processes to observed thermodynamics /." 2001. http://www.library.wisc.edu/databases/connect/dissertations.html.
Повний текст джерела
47
Anderson, Melissa Ann Witmer. "Thermodynamics of the unfolding of a 12 base pair DNA duplex and the interaction of the Laci-DNA binding domain with weak and strong operator DNA." 2008. http://www.library.wisc.edu/databases/connect/dissertations.html.
Повний текст джерела
48
"Investigation on the relationship between structural flexibility and thermodynamics of DNA: insights from NMR structural studies of CODON 335 of HKNPC-EBV LMP1 gene." 2001. http://library.cuhk.edu.hk/record=b6073331.
Повний текст джерелаАнотація:
by Chiu Wing Lok Abe Kurtz.Thesis (Ph.D.)--Chinese University of Hong Kong, 2001.
Includes bibliographical references (p. 218-230).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Mode of access: World Wide Web.
Abstracts in English and Chinese.
49
Nostheide, Sandra. "Kinetics and thermodynamics of unfolding processes in DNA molecules with several conformational states: theory and experiments." Doctoral thesis, 2014. https://repositorium.ub.uni-osnabrueck.de/handle/urn:nbn:de:gbv:700-2014101512898.
Повний текст джерелаАнотація:
The modelling of single-molecule experiments is of vital interest to gain new insights into processes which were hitherto not accessible by measurements performed on bulk systems. In the first part of this thesis, the kinetics of a triple-branch DNA molecule with four conformational states is investigated by employing pulling experiments
with optical tweezers and theoretical modelling. Probability distributions of first rupture forces, which are calculated by
applying transition rate theory to a free energy model, show good agreement with experimental findings. Permanently frayed molecules could be identified by analysing the number of opening base pairs in
force jumps. In the second part of the thesis, DNA hairpin molecules with periodic base sequences are studied. Their unfolding kinetics allows an analytical treatment, because they exhibit a regular coarse-grained free energy landscape as a function of the number of opened base pairs. A procedure is developed for determining all relevant parameters of the landscape, which relies on probabilities that can be
easily sampled from the unfolding trajectories. By means of Monte
Carlo simulations it is shown that already 300 trajectories, as typically measured in single-molecule experiments, provide faithful
results for the energetic parameters. The approach in particular opens a new access to improve loop contributions in the free energy landscape. In the third part of the thesis, a simulation method is developed for
modelling the unfolding kinetics of DNA molecules with arbitrary base sequences. The method is validated against experimental data for five DNA hairpin molecules with different length of the end-loop.
Applications of the method enable one, among others, to improve the parameter determination in functional forms suggested for the tail behaviour of work distributions. Such work distributions enter detailed and integral fluctuation theorems, which are useful for estimating free energy differences between folded and unfolded states from nonequilibrium measurements.
50
Svoboda, Jakub. "Studie strukturních vlastností jednovláknových DNA biofyzikálními metodami a krystalograficky." Master's thesis, 2021. http://www.nusl.cz/ntk/nusl-437785.
Повний текст джерелаАнотація:
DNA is the fundamental molecule in all domains of life, its role in heredity is well established. Although the famous double helical complementary form is indispensable for replication mechanism DNA can occupy wide range of conformations. In the past studies performed in the laboratory, DNA oligomers related to single stranded bacterial Repetitive Extragenic Palindromic (REP) showed spectral behavior suggesting complex equilibria including double helical, hairpin, and tetraplex conformations. The studies presented in this thesis extended the scope of analyzed sequences and employed circular dichroism spectroscopy and X-ray crystallography. We report spectral data and X-ray structures of three successfully crystalized oligonucleotides. All three structures acquire double helical architecture with two consecutive T- T mismatches in the center. To improve the convergence of the refinement process of the crystal structures we used novel dinucleotide conformational classes, NtC classes. The NtC class classification was also used to analyze geometries of selected non-canonical base pairs in all DNA crystal structures in the Protein Data Bank. We measured the fit between geometries of the dinucleotides involved in the non-canonical base pairing and the NtC classes and correlated this fit to the electron...