Teses / dissertações: "Polypeptides"

1

Aronsson, Christopher. "Tunable and modular assembly of polypeptides and polypeptide-hybrid biomaterials". Doctoral thesis, Linköpings universitet, Molekylär fysik, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-132949.

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Biomaterials are materials that are specifically designed to be in contact with biological systems and have for a long time been used in medicine. Examples of biomaterials range from sophisticated prostheses used for replacing outworn body parts to ordinary contact lenses. Currently it is possible to create biomaterials that can e.g. specifically interact with cells or respond to certain stimuli. Peptides, the shorter version of proteins, are excellent molecules for fabrication of such biomaterials. By following and developing design rules it is possible to obtain peptides that can self-assemble into well-defined nanostructures and biomaterials. The aim of this thesis is to create ”smart” and tunable biomaterials by molecular self-assembly using dimerizing –helical polypeptides. Two different, but structurally related, polypeptide-systems have been used in this thesis. The EKIV-polypeptide system was developed in this thesis and consists of four 28-residue polypeptides that can be mixed-and-matched to self-assemble into four different coiled coil heterodimers. The dissociation constant of the different heterodimers range from μM to < nM. Due to the large difference in affinities, the polypeptides are prone to thermodynamic social self-sorting. The JR-polypeptide system, on the other hand, consists of several 42-residue de novo designed helix-loop-helix polypeptides that can dimerize into four-helix bundles. In this work, primarily the glutamic acid-rich polypeptide JR2E has been explored as a component in supramolecular materials. Dimerization was induced by exposing the polypeptide to either Zn2+, acidic conditions or the complementary polypeptide JR2K. By conjugating JR2E to hyaluronic acid and the EKIV-polypeptides to star-shaped poly(ethylene glycol), respectively, highly tunable hydrogels that can be self-assembled in a modular fashion have been created. In addition, self-assembly of spherical superstructures has been investigated and were obtained by linking two thiol-modified JR2E polypeptides via a disulfide bridge in the loop region. The thesis also demonstrates that the polypeptides and the polypeptide-hybrids can be used for encapsulation and release of molecules and nanoparticles. In addition, some of the hydrogels have been explored for 3D cell culture. By using supramolecular interactions combined with bio-orthogonal covalent crosslinking reactions, hydrogels were obtained that enabled facile encapsulation of cells that retained high viability. The results of the work presented in this thesis show that dimerizing α–helical polypeptides can be used to create modular biomaterials with properties that can be tuned by specific molecular interactions. The modularity and the tunable properties of these smart biomaterials are conceptually very interesting andmake them useful in many emerging biomedical applications, such as 3D cell culture, cell therapy, and drug delivery .

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2

Göransson, Ulf. "Macrocyclic polypeptides from plants". Doctoral thesis, Uppsala University, Department of Medicinal Chemistry, 2002. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-1956.

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The aim of this work was to explore the structural and functional diversity of polypeptides that are found in plants. Expanding knowledge of simililarities between plant use of these compound and animal use promises exceptional opportunities for finding, from plant research, new structures with biomedical and biotechnological potential.

A fractionation protocol was developed and applied to many plant species, providing fractions enriched in polypeptides, amenable to chemical and biological evaluation. From one species, the common field pansy (Viola arvensis), a 29-amino-acid residue polypeptide was isolated, named varv A, which revealed a remarkable macrocyclic structure (i.e., N- and C-termini are joined) stabilised by three knotted disulfides.

Varv A, together with an increasing number of homologous peptides, form the currently known peptide family of cyclotides. Their stable structure makes them an attractive scaffold for protein engineering. In addition, they display a wide range of biological activities (e.g., antimicrobial, cytotoxic, and insecticidal). As a part of this work, the cytotoxic effects of varv A and two other isolated cyclotides were evaluated in a human cell-line panel: all were active in the low µM range. Most likely, these effects involve pore formation through cell membranes.

Cyclotides were found to be common in the plant family Violaceae; with eleven cyclotides isolated and sequenced from V. arvensis, V. cotyledon, and Hybanthus parviflorus. For six members of the genus Viola, cyclotide expression profiles were examined by liquid chromatography-mass spectrometry (LC-MS): all expressed notably complex mixtures, with single species containing more than 50 cyclotides. These profiles reflect the evolution of the genus.

To assess these mixtures, a rational strategy for MS based amino acid sequencing of cyclotides was developed, circumventing inherent structural problems, such as low content of positively charged amino acids and the macrocyclic structure. This was achieved by aminoethylation of cysteines, which, following tryptic digestion, produced fragments of size and charge amenable to MS analysis. This method was also modified and used for mapping of disulfide bonds.

Methods for isolation and characterisation developed in this work may prove useful not only for further studies on macrocyclic polypeptides from plants, but also for other plant peptides and disulfide-rich peptides from animals.

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3

Ng, Yuen-lam Stephanie, e 吳宛霖. "Identification of VIP, PACAP and their receptors in agnathans: insights into the ancestral origin of theligands and receptors". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2010. http://hub.hku.hk/bib/B45704879.

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4

Mortenson, Paul Neil. "Energy landscapes of model polypeptides". Thesis, University of Cambridge, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621232.

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5

Monreal, Jorge. "Polymer Characteristics of Polyelectrolyte Polypeptides". Scholar Commons, 2016. http://scholarcommons.usf.edu/etd/6326.

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Polypeptides are polymerized chains of amino acids linked covalently through peptide bonds. Polyelectrolyte polypeptides are polypeptides with electrolyte repeating groups. Several amino acids contain ionizable side chains which result in charge distributions when dissolved in aqueous solutions. This dissertation is motivated by a desire to gain knowledge of polyelectrolyte polypeptides as recent advances in chemical synthesis of polypeptides have made possible the fabrication of designed polypeptides that do not naturally occur in nature. Potential applications of newly designed polypeptides span the range from medical to clothing and energy even to robotics. In this dissertation we compare the characteristic behavior of two polypeptide polyanions: Poly-(L-Glutamic Acid) [PLE] and Poly-(L-Glutamic Acid4, Tyrosine1) [PLEY(4:1)]. Comparative characteristic behaviors of each is conducted through relaxation phenomena in the context of mechanical elasticity measurements of hydrogels and dielectric relaxation of aqueous solutions in a radio frequency range of 1 MHz to 1000 MHz. Hydrogels are fabricated by crosslinking each polyanion with Poly-(L-Lysine) [PLK], a polycation, via the crosslinker 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Elasticity and viscoelasticity measurements are conducted in a fixture designed by our lab. Dielectric relaxation behavior is studied on aqueous solution of both PLEY and PLE using a capacitive fixture, also designed in our lab. RF signals provided by an impedance analyzer are converted to permittivity and dielectric loss measurements. Peaks in dielectric loss provide evidence of relaxation mechanisms. A comparison of experimental results to theoretical expectations reveal both expected and some surprising behavior. Relaxation times for crosslinked hydro-gels scale according to theoretical expectations according to so-called reptation dynamics. However, relaxation times of aqueous solutions did not scale as entangled polyelectrolytes. First, both PLEY and PLE scaled as neutral polymers rather than polyelectrolytes. This was expected because of the high concentrations studied. However, due to the high concentrations, it was expected that polypeptides were entangled in solutions. Data compared to theory did not support this expectation. We, additionally, conducted a self-crosslinking experiment of a polyampholyte: RADA16. RADA16 is known to self-assemble into nano-fibers formed by -sheet stacking. The self-crosslinking was also mediated by EDC. Results of crosslinking showed formation of polypeptide spherules as well as nano-crystals nominally orthorhombic in shape. It was not possible to ascertain composition of the nano-crystals due to both the limited amount of raw material available and the capabilities of measurement equipment as of this writing. It is hypothesized that nano-crystals are composed of some type of urea by-product from the crosslinking reaction. The spherules, on the other hand, seem to be described by the theory of hydrophobic polyelectrolytes. Additional research conducted with regards to electromagnetic hydrodynamic flows during the time frame of this dissertation is also included. The research uses hydrodynamic conservation equations as a starting point to derive one electromagnetic flow momentum equation analogous to the Cauchy momentum equation of hydrodynamics. It also introduces a mass- energy conservation equation for electromagnetic flow that has no hydrodynamic analogue. We begin this dissertation by introducing in Chapter 1 some of the theoretical background necessary to understand results from experiments. Chapter 2 introduces experimental results from elasticity and viscoelasticity measurements and Chapter 3 explains the dielectric relaxation experiment. We then follow with Chapter 4 which presents conclusions from mechanical and dielectric relaxation experiments in a concise format. Results from the self- crosslinking of RADA16 are presented in Chapter 5. Finally, the additional research on electromagnetic flow is presented in Chapter 6.

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6

Crick, F. "Polypeptides and proteins : X-ray studies". Thesis, University of Cambridge, 2013. https://www.repository.cam.ac.uk/handle/1810/250994.

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7

Oomen, Ray. "Interaction of polypeptides with lipid bilayers". Thesis, University of Ottawa (Canada), 1988. http://hdl.handle.net/10393/5253.

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8

Ling, Roger. "Polypeptides of murine and avian pneumoviruses". Thesis, University of Warwick, 1988. http://wrap.warwick.ac.uk/55532/.

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The work described in this thesis identifies some properties of the major polypeptides of pneumonia virus of mice (PVM) and of turkey rhinotracheitis (TRT) virus. The PVM glycoproteins have been studied in particular detail while the results obtained with TRT virus provide a preliminary description of the polypeptides of this virus. Twelve major PVM specific polypeptides designated L, G1, G2, F1, N, 39K, 35K, M, 20K, 19K, 16K and 12K were identified. In addition PVM specific polypeptides designated 25K, 24K, 23K, 18K and 17K were sometimes detected. Monoclonal antibodies directed against the G1/G2, 39K and M polypeptides were produced. The a~ility of a monoclonal antibody to precipitate G1 and G2 suggested that these two glycosylated proteins were related and this was confirmed by tryptic peptide mapping. G2 was shown to be derived from G1 in pulse chase experiments and a similar relationship between two higher mobility polypeptides synthesized in the presence of tunicamycin was observed. The G protein may have a precursor since G1 did not appear immediately following a pulse labelling. The precursor could not however be identified. An additional minor glycosylated polypeptide of 42K was found to be related to the G protein. The F1 protein appeared to be poorly glycosylated and a difference in mobility of the polypeptide synthesized in the presence of tunicamycin did not appear to be directly due to a lack of N-linked oligosaccharides. The polypeptide migrated more slowly under non-reducing conditions but no evidence of a small disulphide bonded polypeptide was found in contrast to the situation with other paramyxoviruses. This polypeptide appeared to be the major PVM protein expressed on the cell surface and was associated with G1 and G2 as the major protein in a particulate fraction of the infected cell supernatant. Tentative relationships were suggested between the 39K, 35K and 25K polypeptides, the M and 24K polypeptides and the 20K and 19K polypeptides. This together with the observation that the 12K polypeptide was not a primary gene product suggested that there may be about 11 PVM polypeptides. The N or 39K and the 20K or 19K polypeptides were observed to be phosphorylated. Twelve possible TRT virus specific polypeptides of 150K, 129K, 95K, 83K, 57K, 45K, 38K, 35K, 3DK, 23K, 19K and 15K were identified. The 150K, 95K, 83K, 57K, 45K and 15K polypeptides were glycosylated with the latter three polypeptides showing a similar relationship to the F1,2, F1 and F2 polypeptides of paramyxoviruses. A broad glycosylated band designated the 31K polypeptide was identified that was similar to a smeared band observed on prolonged exposure of immunoprecipitates of PVM polypeptides labelled with [3H]-glucosamine. The 35K and 19K polypeptides were observed to be phosphorylated. PVM may be more closely related to RS virus than TRT virus since anti-PVM serum irnmunoprecipitated the RS virus N polypeptide but not any TRT virus polypeptides. The PVM 39K polypeptide and the RS virus P protein were recognised by a monoclonal antibody providing further evidence of a relationship between PVM and RS virus.

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9

Galbraith, Toby Patrick. "Biophysical studies of membrane channel polypeptides". Thesis, Birkbeck (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.249171.

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10

Mullin, M. J. "Combinatorial studies on the B-loop residues of human epidermal growth factor". Thesis, Queen's University Belfast, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.273233.

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11

Smith, Dawn. "Protein distribution between cytoplasmic and thylakoid membranes of cyanobacteria". Thesis, University of Cambridge, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.282019.

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12

Deller, Marc Christian. "Structural studies of cytokines and cytokine receptors". Thesis, University of Cambridge, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.326028.

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13

Meillon, Jean Christophe. "Transport transmembranaire et auto-assemblage impliquant des peptides synthétiques modifiés". Sherbrooke : Université de Sherbrooke, 1998.

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14

McColl, Suzzanne May. "Study on polypeptides of cyanobacterial photosystem 2". Thesis, University of Central Lancashire, 1993. http://clok.uclan.ac.uk/21003/.

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The main objective of this work was to characterize cyanobacterial PS2 polypeptides with particular respect to calcium binding. A number of cyanobacteria were studied by trypsin digestion of their thylakoid membranes which showed that using the same method of preparation some cyanobacteria produce everted thylakoid vesicles whilst others produce right sided vesicles. No direct correlation between the sidedness of the produced thylakoids and any growth or metabolic characteristic of the organithm was found. In Synechocystis 6308 a polypeptide of 27kDa , apparently unrelated to known polypeptides of the oxygen evolving complex, was shown to disappear on trypsin treatment, a disappearance which correlated to the loss of oxygen evolving activity. Calcium binding studies performed on Synechococcus 7942 showed the presence of a calmodulin-like calcium binding site within both thylakoids and PS2. The protein on which this site resided was not proven but results pointed to the involvement of the Dl protein. Synechococcus 7942 produces two forms of the Dl protein and mutants in which one form was deleted showed differing affinities for calcium both in vivo (during growth) and in vitro (during reactivation experiments). Differences were also noted for spectra from the mutant strains with one form appearing to retain greater quantities of phycocyanin on the thylakoid membranes. Protein kinase assays performed on the Dl mutants also showed differences between these and the wild type organism. These differences were noted for small polypeptides of 8, 10 and 12kDa. A phosphoprotein of 14kDa was shown to be related to the psb H gene product and a phosphoprotein of approximately 32kDa, possibly Dl, was also obtained. A further 58kDa phosphoprotein which may be related to the calmodulin site or to the protein kinase responsible for PS2 phosphorylation was noted. Both calcium and magnesium were shown to have specific effects on phosphorylation primarily with respect to the small polypeptides mentioned above. Studies on the possible glycosylation of P52 and thylakoid polypeptides produced evidence to suggest that either the Dl or D2 polypeptides are glycosylated and that this may provide a mechanism by which the PS2 complex is held together. Evidence in support of glycosylation of phycobilisome components was also produced with polypeptides of 34.5, 45 78 and 90kDa appearing to be glycosylated. A polypeptide of 50kDa which is closely associated with PS2 was shown to be related , in terms of homology, to a 20kDa DCCD-binding protein of plants which is thought to be a LHC protein.

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15

Wallace, T. Paul. "Extrinsic photosystem II polypeptides in Phormidium laminsoum". Thesis, University of Cambridge, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.335249.

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16

Miramon, Hélène. "Développement de polypeptides hélicoïdaux et activité élicitrice". Montpellier 2, 2009. http://www.theses.fr/2009MON20046.

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Le travail présenté a pour but la synthèse de polypeptides hélicoïdaux par polymérisation de N-carboxyanhydrides d'a-aminoacides. La première partie concerne le développement de polymères éliciteurs, c'est-à-dire capables de stimuler les défenses immunitaires des plantes. Ces polypeptides biodégradables et non toxiques sont constitués d'aminoacides inducteurs d'hélice a, comme l'alanine ou l'acide glutamique. Les polymères obtenus, hydrophobes et insolubles, ont été caractérisés par MALDI-TOF, électrophorèse capillaire de zone, dichroïsme circulaire et RMN. Ces composés ont été testés en utilisant le dosage des peroxidases, marqueur précoce de l'élicitation. La tête de série a ensuite été testée en champ et sa synthèse développée à l'échelle du kilogramme. La deuxième partie de ce travail est dévolue à un analogue non naturel silylé de la proline, la silaproline. La synthèse de ce composé a été optimisée de manière à le produire à plus grande échelle. Puis, les conformations adoptées par la silaproline ont été étudiées dans des dipeptides modèles par modélisation moléculaire, RMN et IR. Enfin, le NCA de la silaproline a été synthétisé dans le but de développer des poly(silaproline)s et d'étudier les conformations adoptées par ces polypeptides
This work is aimed at helical polypeptides synthesis by polymerization of amino-acids N-carboxyanhydrides. The first part concerns the development of elicitor polymers, able to stimulate self defences of plants. These biodegradable and non toxic polypeptides are constituted by amino-acids which induce a-helix, such as alanine and glutamic acid. The hydrophobic and insoluble polymers characterized by MALDI-TOF, zone capillary electrophoresis, circular dichroism and NMR. These compounds have been tested using peroxidase test, early elicitation marker. A leader has been tested in field and its synthesis has been developped to kilogram scale. The second part of this work is devoted to an unnatural sililated analog of proline, the silaproline. The synthesis of this compound have been optimized in order to produce it to large scale. Then, the silaproline conformations have been studied in modeling dipeptides by molecular modelization, NMR and IR. At last, the NCA of silaproline has been synthetized in order to develop poly(silaproline)s and to study the conformations of these polypeptides

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17

Georgilis, Evangelos. "Engineering of Thermoresponsive Diblock Elastin-like Polypeptides". Thesis, Bordeaux, 2019. http://www.theses.fr/2019BORD0444.

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La thèse présentée porte sur l’ingénierie de diblocs de polypeptides à base de motifs élastine (ELPs) susceptibles de s’auto-assembler sous forme de nanoparticules après modification chimique de certains résidus. La stratégie inclue le développement de diblocs d’ELPs composés d’un bloc hydrophobe comportant l’isoleucine en position hôte, fusionné à l’extrémité N-terminale avec un bloc contenant des résidus méthionine chimiosélectivement modifiables. Des modifications du groupement thioéther permettent en effet l’hydrophilisation du segment ELP correspondant ainsi que l’introduction de groupements réactifs. Une première génération d’ELPs diblocs a été développée par génie génétique, production recombinante chez Escherichia coli et caractérisée. Le résidu cystéine à l’extrémité C-terminale a été modifié pour contrôler la monodispersité et introduire des fluorochromes, tandis que les résidus méthionine ont été modifiés pour changer l’équilibre hydrophile/hydrophobe et introduire des groupements réactifs. L’auto-assemblage des diblocs non-modifiés et post-modifiés a été étudiée par des mesures de turbidité et diffusion dynamique de la lumière. Ces méthodes ont mis en évidence une transition thermale de chaînes solubles en agrégats de taille micronique. Pour permettre la formation particules de taille nanométrique, une deuxième génération d’ELPs diblocs a été conçue grâce à l’application d’un modèle empirique. La deuxième génération d’ELPs diblocs forme effectivement des nanoparticules après modification chimiosélective des résidus méthionines. Ces structures pourront potentiellement contribuer au développement de nanoformulations à base d’ELPs
The present work focuses on the engineering of diblock elastin-like polypeptides (ELPs) that thermally assemble into nanostructures after the application of chemical modifications. The strategy involved the development of diblock ELPs composed of a hydrophobic isoleucine-containing block fused at its N-terminal end to a block containing residues amenable to chemoselective modifications, namely methionine. This particular residue was employed because the orthogonal modification of its thioether group allows for the change of the hydrophilic/lipophilic balance of the diblock ELP and the possible simultaneous grafting of functional ligands. A first generation of diblock ELPs was therefore designed by means of molecular clonings, produced in E. coli, and characterized by chemical methods to further monitor post-modifications. The chemical modifications were applied at the C-terminal cysteine to control the system monodispersity and introduce fluorescent probes, and also at methionine in order to change the hydrophilic/lipophilic balance and introduce reactive groups. The self-assembly of the non-modified and post-modified ELPs was monitored by means of turbidimetry, nanoparticle tracking analysis and dynamic light scattering, which showed that these sequences possessed a transition from monomers to aggregates. To access nanoparticle formation, a second generation of diblock ELPs was developed, the design of which was based on theoretical modeling. The second generation diblocks self-assembled into nanoparticles by means of methionine post-modifications. It is expected that these sequences will contribute to the development of diblock ELP-based nano-formulations

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18

Alanazi, Hamdan Noman. "Characterization of Elastin-Like Polypeptides Using Viscometry". Cleveland State University / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=csu1311026986.

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19

Leuthold, Simone Denise. "Structure-function relationship of organic anion transporting polypeptides /". Zürich : ETH, 2007. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=17208.

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20

Turpin, Eleanor R. "Computational studies of folding and binding of polypeptides". Thesis, University of Nottingham, 2013. http://eprints.nottingham.ac.uk/13644/.

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In this thesis molecular dynamics simulations, in conjunction with the complementary methods of docking and QM-MM, are used, and further developed, to study two unusual polypeptide systems: the conformational preferences of isomers of an antibiotic peptide and the binding behaviour of a human transporter protein. The antibiotic peptides are analogues of a naturally occurring antibacterial called nisin which has a biological function dependent on the formation of five macrocyclic rings closed by a thioether bond between modified L-amino and D-amino residues. We propose analogues where the thioether bond is replaced by a disulfide bond between cysteine residues and the chirality of the cysteines is altered. The conformational preferences of the nisin analogues, and the dependence of ring formation on cysteine chirality, are characterised using molecular dynamics. An analogue (D-Cys3-D-Cys7-L-Cys8-L-Cys11) is identified that favours the simultaneous formation of the S3-S7 and S8-S11 disulfide bonds and has an RMSD of 0.6 Å to 1.7 Å between the centroids from clustering the MD trajectories and an NMR structure of wt-nisin. The nisin analogues contain unusual D-amino residues and using explicit solvent MD simulations of four polypeptides, it is shown that the (φ, ψ) → (-φ, -ψ) transformation of the CMAP term in the CHARMM potential energy function leads to sampling of conformations which are closest to X-ray crystallographic structures for D-amino residues and that the standard CMAP correction destabilises D-amino β-sheets and β-turns. The ileal lipid binding protein (ILBP) shows cooperative binding comparable to haemoglobin and unusual site selectivity where one ligand will completely displace another from a binding site, despite both sites having an affinity for each ligand type and the ligands only differing by a single hydroxyl group. A probable location of the third binding site of ILBP is identified which has a role in the allosteric binding mechanism. MD simulations indicate that binding to this exterior site induces changes in the orientation of the α-helices with respect to the β-barrel by ~10°. An energetic mechanism of site selectivity for ILBP is proposed using evidence from MD simulations. The higher hydrophobicity of chenodeoxycholic acid leads it to sit deeper in the binding cavity and interact with Gln-51. This causes the cholic acid ligand to be deeper and induces the helices to move closer to the β-barrel, preventing further ligand exchange.

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21

Gaskell, M. J. "Intracellular fates of microinjected precursor and mature polypeptides". Thesis, University of Nottingham, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.371998.

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22

Ramesh, Balasubramaniyam. "The chemical synthesis and characteriation of biomembrane polypeptides". Thesis, University College London (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.281777.

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23

Nigro, Giuliano. "Incorporation de la beta alanine dans des polypeptides". Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLX064.

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Les cellules vivantes utilisent 20 acides α aminés canoniques lors de la synthèse ribosomale des protéines, même si dans de rares occasions, d’autres acides aminés (selenocysteine ou pyrrolysine) peuvent être utilisés. Le répertoire des acides aminés utilisables est donc assez restreint. Cela limite la possibilité de construire des protéines ayant de nouvelles propriétés. Une tendance forte dans le domaine de l’ingénierie des protéines est la construction de systèmes permettant d’incorporer des acides aminés non canoniques pendant la biosynthèse in vivo. Ces travaux ont connu d’importants succès, mais tous les acides aminés incorporés jusqu'aux années 2010 étaient des acides aminés α. L’incorporation d’acides aminés β à des sites spécifiques créerait une flexibilité supérieure à celle des acides aminés α dans la chaîne principale, ce qui augmenterait les possibilités de repliement de la protéine. Il a été montré récemment en utilisant un système de traduction in vitro de synthèse protéique qu’il était possible d’incorporer des acides aminés β à des positions spécifiques (Katoh and Suga, 2018). Pour parvenir à transposer ce système in vivo, la principale limitation est l’aminoacylation des ARNt avec les acides aminés β. Il a été récemment proposé que certaines aminoacyl-ARNt synthetases étaient capables d’utiliser les acides aminées β, mais cette capacité reste limitée.Le but de cette thèse est d’incorporer la β-méthionine dans des polypeptides in vivo. Pour cela nous avons caractérisé la reconnaissance et l’utilisation de la L-β-homométhionine par la méthionyl-ARNt synthetase (MetRS) d’E. coli. Nous avons en particulier déterminé une structure cristallographique à haute résolution du complexe MetRS: β -Met. En utilisant la spectroscopie de fluorescence ainsi que la spectroscopie de masse, nous avons pu mettre en évidence l’activation de la -Met en adénylate ainsi que son estérification à un ARNtMet. Toutefois, les efficacités mesurées sont très petites par rapport à ce qui avait été publié. Une contamination de la -Met commerciale par de la méthionine pourrait expliquer ces différences. Une étude in silico basée sur la structure du complexe MetRS:β-Met a été menée en collaboration avec l’équipe de Bio-informatique du laboratoire afin de rechercher des enzymes mutantes plus efficaces vis à vis des acides aminés β. Enfin, nous avons amorcé la mise en place d’une méthode d’évolution dirigée destinée à améliorer l’efficacité d’incorporation des acides aminés β in vivo
Living cells use 20 canonical α amino acids during ribosomal protein synthesis, although in rare cases, other amino acids such as selenocysteine or pyrrolysine can be used. The repertoire of usable amino acids is therefore quite limited. This limits the ability to build proteins having new properties. A dominant trend in the field of protein engineering is the construction of systems capable of incorporating non-canonical amino acids during in vivo protein synthesis. These studies have been very successful, but all amino acids incorporated until the 2010s were α-amino acids. Incorporation of β-amino acids at discrete sites would create unprecedented flexibility in the main chain, which would increase the potential for new protein folds. It has recently been shown using an in vitro protein synthesis system that it is possible to incorporate β-amino acids at specific positions (Katoh and Suga, 2018). In order to transpose this system in vivo, the main limitation is the aminoacylation of tRNAs with β -amino acids. It has recently been proposed that some aminoacyl-tRNA synthetases can use β-amino acids, but this capacity remains limited. The aim of this thesis is to incorporate β-methionine in polypeptides in vivo. For this purpose, we have characterized the recognition and use of L-β-homomethionine by methionyl-tRNA synthetase (MetRS) from E. coli. In particular, we have determined a high-resolution crystallographic structure of the MetRS:β-Met complex. Using fluorescence spectroscopy and mass spectroscopy, we were able to demonstrate the activation of -Met into adenylate as well as its esterification onto tRNAMet. However, the measured efficiencies are very small compared to what was published. Contamination of commercial β-Met with methionine may explain these differences. An in silico study based on the structure of the MetRS:β-Met complex was conducted in collaboration with the laboratory's bioinformatics team in order to search for mutant enzymes more efficient in the use of β-amino acids. Finally, we initiated the implementation of a directed evolution method to improve the efficiency of incorporation of amino acids in vivo

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24

Lucas, Paul. "Cationic polypeptides for gene delivery to eukaryotic cells". Thesis, University of Bath, 1995. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.307110.

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Scanlan, D. J. "Biochemical and molecular genetic approaches to studying protein export by cyanobacteria". Thesis, University of Warwick, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.384785.

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Manasse, Robert Samuel. "Photosystem I and cyclic transfer in Phormidium laminosum". Thesis, University of Cambridge, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.336741.

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Petitdemange, Rosine. "Chemoselective modifications of recombinant elastin-like polypeptides : tuning thermosensitivity and bioactivity". Thesis, Bordeaux, 2016. http://www.theses.fr/2016BORD0360/document.

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La thèse présentée porte sur la préparation de dérivés de polypeptides recombinant à base de motifs élastine (ELPs) ainsi que sur l'étude de leurs propriétés physicochimiques et biologiques. Des ELPs contenant des résidus méthionine ont été modifiés de manière chimiosélective soit en utilisant des halogénures d'alkyle ou différents époxydes, soit par oxydation des résidus méthionine. La caractérisation par RMN et par spectrométrie de masse des composés obtenus a permis de confirmer leur fonctionnalisation quantitative. L'étude de la réponse en température de ces dérivés d'ELP par des mesures de turbidité ou par des mesures de diffusion de la lumière a montré le fort impact des modifications entreprises sur la température de transition (TI) des ELPs. Il a également été montré que la n peut être modifiée par échange des contre-ions des dérivés cationiques. Enfin, des monosaccharides ont été conjugué aux ELPs contenant des groupements alcyne par cycloaddition de Huisg en afin d'obtenir des glycopolypeptides. Les propriétés thermosensibles ainsi que les propriétés biologiques de ces conjugués ont été testées et ces dernières ont permis de montrer leur capacité à se lier sélectivement à des lectines. Leur utilisation pour trier des protéines d'intérêt et les redisperser est finalement évaluée de façon préliminaire
This thesis describes the preparation of elastin-like polypeptides (ELPs) derivatives and the study of their physico-chemical and biological properties. Methionine-containing ELPs were chemoselectively modified using either alkyl halides or epoxides or by oxidation of their methionine residues. The successful functionalization was assessed by NMR and mass spectrometry analysis of the resulting compounds. The thermoresponsive properties of these ELP derivatives were evaluated either by light scattering or by turbidity measurements showing the strong effect of these modifications on the ELPs transition temperature (TI). The counterion affect on the thermosensitivity of the polycationic derivatives was also studied. The synthesis of ELP glycopolypeptides was finally achieved by conjugating monosaccharides to the ELP alkyne derivatives through Huisgens cycloaddition. Along with the thermoresponsive properties, the bioactivity of the ELP glycoconjugates was studied and proved their ability to specifically bind lectins. Their use for protein sorting and release was preliminary evidenced

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28

Chittchang, Montakarn Johnston Thomas P. "Effect of secondary structure on paracellular transport of polypeptides". Diss., UMK access, 2004.

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Thesis (Ph. D.)--School of Pharmacy and Dept. of Chemistry. University of Missouri--Kansas City, 2004.
"A dissertation in pharmaceutical sciences and chemistry." Advisor: Thomas P. Johnston. Typescript. Vita. Description based on contents viewed Feb. 23, 2006; title from "catalog record" of the print edition. Includes bibliographical references (leaves 202-223). Online version of the print edition.

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Stark, Margareta. "Isolation and characterization of lipid-associated and neurosecretory polypeptides /". Stockholm, 2000. http://diss.kib.ki.se/2000/91-628-4202-1/.

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30

Leiper, Kenneth Alexander. "Beer polypeptides and their selective removal with silica gels". Thesis, Heriot-Watt University, 2002. http://hdl.handle.net/10399/444.

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Flint, Daniel Geoffrey. "A molecular modelling study of covalently cross-linked polypeptides". Thesis, University of Birmingham, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.411850.

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Ray, Nicola. "Genetic manipulation of photosystem two polypeptides in Chlamydomonas reinhardtii". Thesis, University College London (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.395045.

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Bishop, Cleo Lucinda. "An investigation of the small polypeptides of photosystem ll". Thesis, University College London (University of London), 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.271720.

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Engler, Amanda Catherine. "Synthesis and testing of novel polypeptides for biological applications". Thesis, Massachusetts Institute of Technology, 2011. http://hdl.handle.net/1721.1/62732.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, 2011.
Cataloged from PDF version of thesis.
Includes bibliographical references.
Natural systems produce macromolecules that assemble into complex, highly ordered structures. In particular, proteins and peptides derived from the 20 naturally occurring amino acids are sequenced macromolecules that can fold into secondary and tertiary structures and can self assemble into quaternary structures. Through weak interactions, these ordered systems produce high-strength materials, provide physical cues to induce cell functions and morphologies, efficiently harvest energy, and transport materials. One of the key challenges in the field of polymer chemistry is the ability to generate synthetic systems that can demonstrate the highly ordered structure, self-assembly, and responsive behavior of these macromolecules. Synthetic polypeptides have received attention because of their unique structural properties and biocompatibility. Like their naturally occurring analogs, these molecules have a poly(amino acid) backbone and posses the ability to fold into secondary structures. Synthetic homo polypeptides are synthesized by the ring opening polymerization of N-carboxyanhydrides formed from naturally occurring amino acids. Although these macromolecules' secondary structure can be controlled to some extent, we are limited by the given side chain, which dictates polymer function, structure, and responsive behavior to temperature or pH among many other properties. We have developed a new approach to the manipulation of synthetic polypeptide composition and function through the introduction of a new NCA polymer, poly(Y-propargyl-L-glutamate) (PPLG) which contains a pendant alkyne group that can be reacted with an azide by the 1,3 cycloaddition "click" reaction. With this system, we can incorporate functional groups that are ordinarily difficult to introduce because of cross-reactions or exhaustive protection-deprotection steps. In addition, we can more directly mimic the adaptive function and responsive behavior of naturally occurring polypeptides. This thesis focuses on the development of the PPLG system and the use of the system for synthetic biomimics, drug delivery, and gene delivery. For synthetic biomimics, as an initial example, densely grafted polymers were synthesized to demonstrate the utility of this synthetic approach. In addition, synthetic antimicrobial polypeptides were synthesized to mimic naturally occurring antimicrobial peptides. For drug and gene delivery, a library of pH responsive peptides were synthesized and characterized.
by Amanda Catherine Engler.
Ph.D.

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35

Morey, Shannon Marie. "Development and study of synthetic polypeptides for biomaterial applications". Thesis, Massachusetts Institute of Technology, 2013. http://hdl.handle.net/1721.1/82333.

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Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2013.
Vita. Cataloged from PDF version of thesis.
Includes bibliographical references (p. 38-39).
Creating new scaffolds for cells is critical to the development of new tissue engineering techniques. In this work, the synthesis of new polypeptide systems is discussed. These systems are intended for the formation of hydrogels which can then be used as cell substrates. Attempts at using the clickable synthetic polypeptide poly(ypropargyl L-glutamate) (PPLG) to form a self-assembly amphiphilic system is discussed, as is the formation of potentially amphiphilic block copolymers with PPLG. The synthesis of a hydrolytically stable synthetic polypeptide with click functionality is also investigated. Additionally, the creation of a polypeptide system with two functionalities available for orthogonal click chemistry is discussed.
by Shannon Marie Morey.
S.M.

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36

Zholobko, Oksana. "Functional Colloids from Amphiphilic Polymer Assemblies and Peptides/Polypeptides". Diss., North Dakota State University, 2019. https://hdl.handle.net/10365/29963.

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The use of responsive polymers, where even minor changes in one of the macromolecular characteristics triggered by the external stimuli can cause drastic changes in the material function or performance, is widely studying area of research. Formation of the thermodynamically stable polymer-peptide colloids, such as mixed micellar assemblies or polymer-enzyme conjugates, loading capacity of the colloids, and cargo activity all depend on the macromolecular interactions within the peptide/polypeptide-polymer system. The goal of this work is to investigate interactions between range of new polymers and various cargo molecules and determine whether those interactions affect the physicochemical properties of the resulted colloids. For this purpose, two types of colloid systems were explored: i) peptide-loaded invertible micellar assemblies (IMAs), formed using hydrophobic interactions between amphiphilic invertible polymers (AIP) and peptides (HA, V5, or peptide-based vaccine), and ii) polymeric cellulosomes made from polymer ligand (PL), copolymer of glycidyl methacrylate (GMA) and poly(ethylene glycol) methyl ether methacrylate (PEGMA) and mixture of cellulases, using covalent bonding. The purpose of the research was to evaluate if colloids properties are affected by changes in responsive polymer characteristics as well as if the developed macromolecular structure and composition need further synthetic modification/optimization. AIP-related part of this dissertation is focused on i) understanding of interaction between peptides and AIPs, and formation of mixed micellar assemblies; ii) further behavior of cargo peptide molecules in the micellar interior under the AIP conformational changes, triggered by IMAs localization at polar and nonpolar interface; iii) evaluation of the impact of IMAs on model lipid membrane diffusivity and permeability. Besides, AIP-peptide assemblies were tested in vitro and in vivo in order to evaluate the cargo delivery, antibody response, and immunity protection in vaccinated pigs against Swine influenza viruses (SIV). To explore the feasibility of covalent bonding in formation of responsive polymer-based colloids, enzyme-polymer conjugates (EPCs) were designed and their enzymatic catalytic activity for the biomass hydrolysis was further tested. The effect of conjugation on catalytic activity, conjugation efficiency, glucose inhibition effect, type of substrate, and type of biomass pretreatment were evaluated and compared to free enzymes.
North Dakota. Department of Commerce
National Science Foundation (U.S.)

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37

Agut, Willy. "Conception de nano-objets adaptatifs à base de polypeptides". Thesis, Bordeaux 1, 2008. http://www.theses.fr/2008BOR12319/document.

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Les copolymères à blocs représentent de bons candidats pour des applications biomédicales, notamment pour l’encapsulation et le relargage contrôlé de médicaments. Depuis quelques années, les équipes de recherche tentent de concevoir des nanoparticules fonctionnelles « sur mesure » de façon à augmenter leur efficacité en tant que nano-vecteurs. C’est dans ce contexte que s’est s’inscrit ce travail dont le but est de concevoir des copolymères à blocs multi-stimulables en vue d’obtenir des nano-objets adaptatifs en milieu aqueux, capables d’encapsuler des molécules actives et de les relarguer de manière contrôlée en jouant sur différents paramètres extérieurs (pH, T...). Ce manuscrit traite en premier lieu d’une nouvelle stratégie de synthèse des copolymères à blocs stimulables “hybrides”, c’est à dire comprenant un bloc vinylique stimulable et un bloc peptidique. Celle-ci, fondée sur le couplage par « chimie click » d’homopolymères fonctionnalisés eux-mêmes obtenus par polymérisation « vivante / contrôlée », s’est révélée très efficace pour l’obtention de copolymères de structure moléculaire très bien définie. . L'autre partie, dédiée à l'étude complète du comportement en milieu aqueux des copolymères à blocs “hybrides”, en fonction de divers paramètres extérieurs (pH, T), illustre leurs propriétés singulières d’auto-assemblage. Ils forment, en effet, une multitude de morphologies différentes mises en évidence par des techniques de caractérisation complémentaires. Cette partie traite également de l’incorporation de particules magnétiques au sein des nanoparticules polymères, afin de concevoir des nano-vecteurs utilisables pour l’imagerie médicale ou pour leurs propriétés d’hyperthermie. Ce projet ambitieux, croisant les concepts de l’auto-assemblage, de l’ingénierie macromoléculaire, de l’encapsulation, des systèmes hybrides organique-inorganique et des systèmes stimulables, constitue une thématique de recherche novatrice
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38

Hayward, Richard Laurence. "Inelastic neutron scattering spectroscopy of polypeptides and molecular crystals". Thesis, University of Edinburgh, 1995. http://hdl.handle.net/1842/21296.

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A predictive and practical theory for a fundamental biological problem - the relation between a protein's three dimensional folded form and its function - will rest on an accurate description of the potential energy surface as a function of the protein configuration, and, thereby, on an accurate description of dynamic and thermodynamic properties. A successful theory of this sort will provide the means for rational design of proteins and ligands with desirable properties. Modern computational chemistry techniques have been applied, with qualitative success, to the calculation of protein potential energy functions, and resulting dynamics and ligand binding properties. These calculations have led to suggestions for new drug design. Improvement in the accuracy of predictions from such calculations will require a consistent program of refinement of parameterisation and approximation schemes, by comparison with experimental data. This thesis describes the application to this task of inelastic neutron scattering experiments on samples of polypeptides (collagen, (prolylprolylglycine)₁₀ and polyproline II) and molecular crystals of biological relevance (acetanilide and two isotopmers). The experimental data were analysed for each of the samples in the context of models for their dynamics on the picosecond time scale. Improvements of the form and parameters of the dynamical models are suggested by comparison of the experimental data with the results of numerical calculations. An appendix described an idea I have had for the model independent exploitation of neutron scattering data, and a second appendix records inelastic neutron scattering data collected for two further molecular crystals of biological relevance, 1-alanine and acetyl-alanyl-methylamide.

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39

Ma, Jiao. "Discovery and Characterization of Novel smORF-Encoded Polypeptides (SEPs)". Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:26718718.

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Peptides and small proteins have essential physiological roles including metabolism (insulin), sleep (orexin), and stress (corticotropin-releasing hormone). Recent exploration of the human genome and proteome has revealed the existence of hundreds to thousands of short open reading frames (sORFs); however, the extent to which sORFs are translated into polypeptides is unknown. Inline with the current convention, a protein-coding short ORF is defined to be a small ORF or smORF; the protein product as a smORF-encoded polypeptide is called a SEP; and a sORF or smORF upstream from an open reading frame (i.e. in the 5’-UTR) is called an upstream ORF or uORF. The identification of smORFs and SEPs have prompted efforts determine the regulation and biological functions for these molecules. My thesis research focused on improving SEP discovery and the characterization of functional SEPs. The discovery of novel SEPs contributes to our understanding of composition of the human genome and proteome. My colleagues and I developed and utilized a proteogenomics strategy, which integrates genomics (RNA-Seq) with proteomics, to discover 86 novel human SEPs, the largest number of validated SEPs described at the time. Our findings indicated that SEPs are a large, unappreciated, peptide family. Moreover, our approach was far from optimized and we felt that there were likely many additional SEPs in the human genome. One goal of my thesis work was to improve the SEP discovery methodology to find more human SEPs. My efforts led to the discovery of an over 300 SEPs in cell lines and human tissue. A second goal of my thesis work was to identify and characterize functional SEPs. To do this I identified the SEPs that are most highly conserved throughout evolution with a program called PhyloCSF. PhyloCSF identifies which SEPs are evolutionary conserved to provide evidence for function. Seven out of the 300 plus SEPs had PhyloCSF scores that indicate that they have been conserved throughout evolution. These seven SEPs included an interesting SEP called SLC35A4-SEP that is generated from a uORF in the SLC35A4 gene. The SLC35A4-SEP had contained a transmembrane domain and analysis of cells revealed the mitochondrial localization of this SEP. Further characterization of SCL35A4 indicated that this polypeptide interacts with members of the ATP synthase complex. Though this interaction requires further validation the putative interactions suggested a role for SLC35A4-SEP in cellular energetics. Overexpression or knockout of SLC35A4-SEP affected cellular respiration. Ongoing work is testing to see if SLC35A4-SEP also effects mitochondrial membrane potential and structure of ATP-synthase. More generally, this approach highlights how I can begin to identify functional SEPs using a combination of computational and experimental methods. And my work on another functional SEP called NoBody indicates that this strategy is general.
Chemistry and Chemical Biology

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40

Watson, David C. (David Charles) Carleton University Dissertation Biology. "Analysis of polypeptides associated with Typula iridescent virus particles". Ottawa, 1993.

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41

Agut, Willy Lecommandoux Sébastien Taton Daniel. "Conception de nano-objets adaptatifs à base de polypeptides". S. l. : Bordeaux 1, 2008. http://ori-oai.u-bordeaux1.fr/pdf/2008/AGUT_WILLY_2008.pdf.

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42

Dauber-Osguthorpe, Pnina. "Conformation and internal motion of polypeptides : molecular dynamics simulations". Thesis, University of Bath, 1990. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.252972.

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43

Pakstis, Lisa M. "Controlled self-assembly of amphiphilic diblock copolypeptides". Access to citation, abstract and download form provided by ProQuest Information and Learning Company; downloadable PDF file 14.66 Mb., 139 p, 2006. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:3200558.

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44

Gao, Yi. "Development of a novel hTERTC27 based cancer : gene therapy /". Click to view the E-thesis via HKUTO, 2007. http://sunzi.lib.hku.hk/HKUTO/record/B39557790.

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45

Wright, Penelope A. "Mechanistic studies on the catalysis and inhibition of serine proteases". Thesis, University of Oxford, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.302492.

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46

Qiu, Guang, e 邱光. "Assessment of the role of corticosterone and adiponectin in the neuroprotective effect of dietary restriction". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2008. http://hub.hku.hk/bib/B41290604.

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Qiu, Guang. "Assessment of the role of corticosterone and adiponectin in the neuroprotective effect of dietary restriction". Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B41290604.

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48

Cowie, David. "Differential induction of organic anion transporting polypeptides in rat liver". Thesis, Available from the University of Aberdeen Library and Historic Collections Digital Resources. Restricted: no access until April 24, 2020, 2008. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?application=DIGITOOL-3&owner=resourcediscovery&custom_att_2=simple_viewer&pid=25707.

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49

Lundh, Ann-Christin. "The rational design of catalytically active polypeptides with supersecondary structure". Göteborg : Department of Organic Chemistry, University of Göteborg, 1995. http://catalog.hathitrust.org/api/volumes/oclc/39066015.html.

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50

Fesinmeyer, Robert Matthew. "Chemical shifts define the structure and folding thermodynamics of polypeptides /". Thesis, Connect to this title online; UW restricted, 2005. http://hdl.handle.net/1773/11621.

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