Literatura académica sobre el tema "NMR Biomolecular structure Interactions and Dynamic"

In recent years, a dynamic view of the structure and function of biological macromolecules is emerging, highlighting an essential role of dynamic conformational equilibria to understand molecular mechanisms of biological functions. The structure of a biomolecule, i.e. protein or nucleic acid in solution, is often best described as a dynamic ensemble of conformations, rather than a single structural state. Strikingly, the molecular interactions and functions of the biological macromolecule can then involve a shift between conformations that pre-exist in such an ensemble. Upon external cues, such population shifts of pre-existing conformations allow gradually relaying the signal to the downstream biological events. An inherent feature of this principle is conformational dynamics, where intrinsically disordered regions often play important roles to modulate the conformational ensemble. Unequivocally, solution-state NMR spectroscopy is a powerful technique to study the structure and dynamics of such biomolecules in solution. NMR is increasingly combined with complementary techniques, including fluorescence spectroscopy and small angle scattering. The combination of these techniques provides complementary information about the conformation and dynamics in solution and thus affords a comprehensive description of biomolecular functions and regulations. Here, we illustrate how an integrated approach combining complementary techniques can assess the structure and dynamics of proteins and protein complexes in solution.

Solid-state nuclear magnetic resonance (ssNMR) is an indispensable tool for elucidating the structure and dynamics of insoluble and non-crystalline biomolecules. The recent advances in the sensitivity-enhancing technique magic-angle spinning dynamic nuclear polarization (MAS-DNP) have substantially expanded the territory of ssNMR investigations and enabled the detection of polymer interfaces in a cellular environment. This article highlights the emerging MAS-DNP approaches and their applications to the analysis of biomolecular composites and intact cells to determine the folding pathway and ligand binding of proteins, the structural polymorphism of low-populated biopolymers, as well as the physical interactions between carbohydrates, proteins, and lignin. These structural features provide an atomic-level understanding of many cellular processes, promoting the development of better biomaterials and inhibitors. It is anticipated that the capabilities of MAS-DNP in biomolecular and biomaterial research will be further enlarged by the rapid development of instrumentation and methodology.

Oxygen is a key atom that maintains biomolecular structures, regulates various physiological processes, and mediates various biomolecular interactions. Oxygen-17 (17O), therefore, has been proposed as a useful probe that can provide detailed information about various physicochemical features of proteins. This is attributed to the facts that (1) 17O is an active isotope for nuclear magnetic resonance (NMR) spectroscopic approaches; (2) NMR spectroscopy is one of the most suitable tools for characterizing the structural and dynamical features of biomolecules under native-like conditions; and (3) oxygen atoms are frequently involved in essential hydrogen bonds for the structural and functional integrity of proteins or related biomolecules. Although 17O NMR spectroscopic investigations of biomolecules have been considerably hampered due to low natural abundance and the quadruple characteristics of the 17O nucleus, recent theoretical and technical developments have revolutionized this methodology to be optimally poised as a unique and widely applicable tool for determining protein structure and dynamics. In this review, we recapitulate recent developments in 17O NMR spectroscopy to characterize protein structure and folding. In addition, we discuss the highly promising advantages of this methodology over other techniques and explain why further technical and experimental advancements are highly desired.

In-cell nuclear magnetic resonance (NMR) spectroscopy offers the possibility to study proteins and other biomolecules at atomic resolution directly in cells. As such, it provides compelling means to complement existing tools in cellular structural biology. Given the dominance of electron microscopy (EM)-based methods in current structure determination routines, I share my personal view about the role of biomolecular NMR spectroscopy in the aftermath of the revolution in resolution. Specifically, I focus on spin-off applications that in-cell NMR has helped to develop and how they may provide broader and more generally applicable routes for future NMR investigations. I discuss the use of ‘static’ and time-resolved solution NMR spectroscopy to detect post-translational protein modifications (PTMs) and to investigate structural consequences that occur in their response. I argue that available examples vindicate the need for collective and systematic efforts to determine post-translationally modified protein structures in the future. Furthermore, I explain my reasoning behind a Quinary Structure Assessment (QSA) initiative to interrogate cellular effects on protein dynamics and transient interactions present in physiological environments.

Various recent developments in solid-state nuclear magnetic resonance (ssNMR) spectroscopy have enabled an array of new insights regarding the structure, dynamics, and interactions of biomolecules. In the ever more integrated world of structural biology, ssNMR studies provide structural and dynamic information that is complementary to the data accessible by other means. ssNMR enables the study of samples lacking a crystalline lattice, featuring static as well as dynamic disorder, and does so independent of higher-order symmetry. The present study surveys recent applications of biomolecular ssNMR and examines how this technique is increasingly integrated with other structural biology techniques, such as (cryo) electron microscopy, solution-state NMR, and X-ray crystallography. Traditional ssNMR targets include lipid bilayer membranes and membrane proteins in a lipid bilayer environment. Another classic application has been in the area of protein misfolding and aggregation disorders, where ssNMR has provided essential structural data on oligomers and amyloid fibril aggregates. More recently, the application of ssNMR has expanded to a growing array of biological assemblies, ranging from non-amyloid protein aggregates, protein–protein complexes, viral capsids, and many others. Across these areas, multidimensional magic angle spinning (MAS) ssNMR has, in the last decade, revealed three-dimensional structures, including many that had been inaccessible by other structural biology techniques. Equally important insights in structural and molecular biology derive from the ability of MAS ssNMR to probe information beyond comprehensive protein structures, such as dynamics, solvent exposure, protein–protein interfaces, and substrate–enzyme interactions.

Nearly all retroviruses selectively package two copies of their unspliced RNA genomes from a cellular milieu that contains a substantial excess of non-viral and spliced viral RNAs. Over the past four decades, combinations of genetic experiments, phylogenetic analyses, nucleotide accessibility mapping, in silico RNA structure predictions, and biophysical experiments were employed to understand how retroviral genomes are selected for packaging. Genetic studies provided early clues regarding the protein and RNA elements required for packaging, and nucleotide accessibility mapping experiments provided insights into the secondary structures of functionally important elements in the genome. Three-dimensional structural determinants of packaging were primarily derived by nuclear magnetic resonance (NMR) spectroscopy. A key advantage of NMR, relative to other methods for determining biomolecular structure (such as X-ray crystallography), is that it is well suited for studies of conformationally dynamic and heterogeneous systems—a hallmark of the retrovirus packaging machinery. Here, we review advances in understanding of the structures, dynamics, and interactions of the proteins and RNA elements involved in retroviral genome selection and packaging that are facilitated by NMR.

The role of biomolecular condensates in regulating biological function and the importance of dynamic interactions involving intrinsically disordered protein regions (IDRs) in their assembly are increasingly appreciated. While computational and theoretical approaches have provided significant insights into IDR phase behavior, establishing the critical interactions that govern condensation with atomic resolution through experiment is more difficult, given the lack of applicability of standard structural biological tools to study these highly dynamic large-scale associated states. NMR can be a valuable method, but the dynamic and viscous nature of condensed IDRs presents challenges. Using the C-terminal IDR (607 to 709) of CAPRIN1, an RNA-binding protein found in stress granules, P bodies, and messenger RNA transport granules, we have developed and applied a variety of NMR methods for studies of condensed IDR states to provide insights into interactions driving and modulating phase separation. We identify ATP interactions with CAPRIN1 that can enhance or reduce phase separation. We also quantify specific side-chain and backbone interactions within condensed CAPRIN1 that define critical sequences for phase separation and that are reduced by O-GlcNAcylation known to occur during cell cycle and stress. This expanded NMR toolkit that has been developed for characterizing IDR condensates has generated detailed interaction information relevant for understanding CAPRIN1 biology and informing general models of phase separation, with significant potential future applications to illuminate dynamic structure–function relationships in other biological condensates.

In this review on advanced biomolecular EPR spectroscopy, which addresses both the EPR and NMR communities, considerable emphasis is put on delineating the complementarity of NMR and EPR regarding the measurement of interactions and dynamics of large molecules embedded in fluid-solution or solid-state environments. Our focus is on the characterization of protein structure, dynamics and interactions, using sophisticated EPR spectroscopy methods. New developments in pulsed microwave and sweepable cryomagnet technology as well as ultrafast electronics for signal data handling and processing have pushed the limits of EPR spectroscopy to new horizons reaching millimeter and sub-millimeter wavelengths and 15 T Zeeman fields. Expanding traditional applications to paramagnetic systems, spin-labeling of biomolecules has become a mainstream multifrequency approach in EPR spectroscopy. In the high-frequency/high-field EPR region, sub-micromolar concentrations of nitroxide spin-labeled molecules are now sufficient to characterize reaction intermediates of complex biomolecular processes. This offers promising analytical applications in biochemistry and molecular biology where sample material is often difficult to prepare in sufficient concentration for NMR characterization. For multifrequency EPR experiments on frozen solutions typical sample volumes are of the order of 250 μL (S-band), 150 μL (X-band), 10 μL (Q-band) and 1 μL (W-band). These are orders of magnitude smaller than the sample volumes required for modern liquid- or solid-state NMR spectroscopy. An important additional advantage of EPR over NMR is the ability to detect and characterize even short-lived paramagnetic reaction intermediates (down to a lifetime of a few ns). Electron–nuclear and electron–electron double-resonance techniques such as electron–nuclear double resonance (ENDOR), ELDOR-detected NMR, PELDOR (DEER) further improve the spectroscopic selectivity for the various magnetic interactions and their evolution in the frequency and time domains. PELDOR techniques applied to frozen-solution samples of doubly spin-labeled proteins allow for molecular distance measurements ranging up to about 100 Å. For disordered frozen-solution samples high-field EPR spectroscopy allows greatly improved orientational selection of the molecules within the laboratory axes reference system by means of the anisotropic electron Zeeman interaction. Single-crystal resolution is approached at the canonical g-tensor orientations—even for molecules with very small g-anisotropies. Unique structural, functional, and dynamic information about molecular systems is thus revealed that can hardly be obtained by other analytical techniques. On the other hand, the limitation to systems with unpaired electrons means that EPR is less widely used than NMR. However, this limitation also means that EPR offers greater specificity, since ordinary chemical solvents and matrices do not give rise to EPR in contrast to NMR spectra. Thus, multifrequency EPR spectroscopy plays an important role in better understanding paramagnetic species such as organic and inorganic radicals, transition metal complexes as found in many catalysts or metalloenzymes, transient species such as light-generated spin-correlated radical pairs and triplets occurring in protein complexes of photosynthetic reaction centers, electron-transfer relays, etc. Special attention is drawn to high-field EPR experiments on photosynthetic reaction centers embedded in specific sugar matrices that enable organisms to survive extreme dryness and heat stress by adopting an anhydrobiotic state. After a more general overview on methods and applications of advanced multifrequency EPR spectroscopy, a few representative examples are reviewed to some detail in two Case Studies: (I) High-field ELDOR-detected NMR (EDNMR) as a general method for electron–nuclear hyperfine spectroscopy of nitroxide radical and transition metal containing systems; (II) High-field ENDOR and EDNMR studies of the Oxygen Evolving Complex (OEC) in Photosystem II, which performs water oxidation in photosynthesis, i.e., the light-driven splitting of water into its elemental constituents, which is one of the most important chemical reactions on Earth.

Diss. (sammanfattning) Stockholm : Stockholms universitet, 2009.
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Submitted. Paper 4: Submitted. Paper 5: In progress. Paper 6: In progress. Paper 7: Manuscript. Härtill 7 uppsatser.

Understanding the energetic and dynamic behavior of natural protein fluctuations is critical to elucidating important information associated with a multitude of protein functions including signaling processes, enzyme behavior, aggregation pathways etc... This information is also critically important in the development of novel and effective strategies aimed at target proteins associated with pathologies and disease. In order to obtain such useful information, tools and techniques are lacking that: 1) permit the efficient quantitative analysis of fluctuation behavior of existing protein structure ensembles and 2) permit computationally generated natural fluctuation states of proteins at relatively large timescales demanded by the need for biologically relevant results. This thesis presents such methods as well as the results of their application to a case study of Aβ40 and pathogenic Aβ42 where we identify key differences in energy interactions between those two isoforms. Additionally, our detailed atom-level analysis, was able to identify very minute differences in Ramachandran angles between the two strains as the cause for these interaction energy differences. We also demonstrate the efficacy of our implicit solvent algorithm in recovering independently, experimentally identified domain motion over a variety of protein systems. Such a system that is medically significant is the HIV-1 protease for which we identified significant motion of a flap domain known to be pharmaceutically important to the protease’s active site in drug targeting strategies. Lastly, we employ the insights thus acquired from the Aβ40/42 case study to see if Aβ42 aggregation inhibitors can be rationally developed and then tested in vitro for their efficacy. Results were very promising with Aβ42 aggregate sizes being significantly reduced by statistically significant margins by the inhibitor compounds. Due to these encouraging results, we have consequently obtained a provisional patent application for our inhibitors.

HU est une protéine bactérienne qui est impliquée dans de nombreuses fonctions liées à l'ADN. Elle est présente sous forme de trois dimères chez E. coli (deux homodimères et un hétérodimère). Lorsque les deux homodimères sont mélangés in vitro, ils échangent leurs chaînes pour former l'hétérodimère. Mon travail a consisté à caractériser, structuralement et cinétiquement, ce mécanisme d'échange qui peut être décrit comme une réaction d’ordre 2 se déroulant en 3 étapes : d'une conformation native (N2) de chaque homodimère à une conformation intermédiaire (I2, partiellement dissociée et déstructurée), puis la formation d'un tétramère transitoire (étape limitante) qui se dissocie finalement en deux hétérodimères. Les résidus considérés comme étant les déterminants structuraux permettant la transition entre N2 et I2 ont pu être déterminés. Ces résidus, enfouis dans N2, forment un patch hydrophobe sur la surface de I2. Ce patch peut être impliqué dans la reconnaissance des chaînes de HU et permettrait la formation du tétramère. MC1 participe à l'organisation du génome de plusieurs archées, à la transcription de l'ADN et à la division cellulaire par des mécanismes inconnus. Nous présentons la structure d'un complexe formé par MC1 avec un ADN de 15 paires de bases. Alors que la protéine a besoin d'adapter sa conformation légèrement, l'ADN subit une courbure dramatique et une torsion impressionnante. Une telle conformation en V du complexe et un modèle structural de MC1 avec un ADN plus long nous ont amené à proposer un nouveau mode de liaison de la protéine en tant qu’« enrouleur ». Des expériences de diffraction RX et de SAXS ont été réalisées sur ce complexe. Malheureusement, la structure n'a pas pu être résolue en raison du manque de données de diffraction et les données SAXS ont invalidé le modèle. Ces résultats confirment que MC1 est une protéine atypique, qui stabilise de multiples conformations en V de l'ADN de manière flexible et dynamique
HU is an essential bacterial protein that is involved in many functions related to DNA. It is present as three dimers in E.coli (two homodimers and one heterodimer). If the two homodimers are mixed in vitro, they exchange their chains to spontaneously form the heterodimer. My work was to characterize, structurally and kinetically, this exchange mechanism that can be described as a second order reaction of three successive steps : from a native conformation of each homodimer into an intermediate homodimer conformation (partially unfolded and dissociated), followed by the formation of a transient tetramer (limiting step) which finally dissociates into two heterodimers. The key residues allowing the protein to switch from the native to the intermediate state has been determined. Whereas, there are buried in the native conformation, they are forming a hydrophobic patch at the surface of the intermediate one. This patch could mediate the association of the intermediate conformation in order to form the tetramer.MC1 participates in the genome organization of several archaea and in DNA transcription and cellular division through unknown mechanisms. We discuss the solution structure of a complex formed by MC1 with a strongly distorted 15 base pairs DNA. While the protein just needs to adapt slightly its conformation, the DNA undergoes a dramatic curvature and an impressive torsion. Such a V-turn conformation of the complex lead us to propose a new binding mode for the protein as a wrapper and a structural model of MC1 with a longer DNA. XR diffraction and SAXS experiments were then carried out on this new complex. Unfortunately, the structure could not be solved due to the lack of diffraction data and the SAXS data invalidated the model. These results confirm that MC1 is an atypical protein, which stabilizes multiple V-turn conformations of the DNA in a flexible and dynamic manner

My thesis is divided into five chapters, starting with a general introduction in first chapter and sample preparation and protein-NMR assignment techniques in second chapter. The remaining three chapters focus on three different areas/projects that I have worked on. Chapter 1: Introduction to nanomaterials and all the experimental techniques This chapter reviews different kinds of nanomaterials and their application utilized for protein-nanomaterial interaction in our study, along with the introduction to different spectroscopy and microscopy techniques used for the interaction studies. Starting with introduction of nanomaterials and all the experimental techniques, which constitute the arsenal for structural studies of the protein-nanomaterial interaction, different steps enroute to structural and dynamic interaction are outlined in detail. Chapter 2: Preparation and Characterization of Proteins used for nanomaterial interaction studies Proteins are generally of three kinds- globular (structured), intrinsically disordered and membrane bound. These proteins have different functions in living organisms and play a major role to maintain metabolism and other important factors. To probe protein-nanomaterial interactions, we have chosen different protein/peptides. This chapter describes the protocol/procedure used for purifying the proteins. For studying a globular protein, ubiquitin was chosen. Nanomaterial-IDP interaction was investigated using the intrinsically disordered central linker domain of human insulin like growth factor binding protein-2 (L-hIGFBP2). The hydrophobic membrane interacting part of the prion protein was chosen as a representative membrane protein. The characterization of the proteins by NMR spectroscopy is also described. Chapter 3: A nanomaterial based novel macromolecular crowding agent Carbon quantum dots (CQD) are nanomaterials with size less than 10 nm, first obtained in 2004 during purification of single-walled carbon-nanotubes. Since then CQDs have been used in a wide range of applications due to their low cost of preparation and favorable properties such as chemical inertness, biocompatibility, non-toxicity and solubility in aqueous medium. One of the applications of CQDs has been their use for imaging and tracking proteins inside cells, based on their intrinsic fluorescence. Further, quantum dots exhibit concentration dependent aggregation while retaining their solubility. Fluorescent carbon quantum dots (CQD) induce macromolecular crowding making them suitable for probing the structure, function and dynamics of both hydrophilic and hydrophobic peptides/ proteins under near in-cell conditions. We have prepared hydrophilic and hydrophobic quantum dots to see the crowding effect. After characterization of CQD, we tested the property of proteins with CQD and found that CQD behaves as a macromolecular crowding agent by mimicking near in-cell conditions. In our study, we have chosen a globular protein, an intrinsically disordered protein (IDP) and one hydrophobic membrane peptide. We have also compared the crowding property of CQD with ficoll which is widely used commercial crowding agent. The overall study tells that the CQD acts like crowding agent and can be used for the study of macromolecular crowding effect. This makes them suitable for structural and functional studies of proteins in near in-cell conditions. Chapter 4: Ubiquitin-Graphene oxide interactions Described here is the interaction of human ubiquitin with GO using NMR spectroscopy and other techniques such as Fluorescence spectroscopy, isothermal titration calorimetry (ITC), UV-Visible spectroscopy, dynamic light scattering (DLS), zeta potential measurements and transmission electron microscopy (TEM). The globular protein ubiquitin interacts with GO and undergoes a dynamic and reversible association-dissociation in a fast exchange regimen as revealed by NMR spectroscopy. The conformation of the protein is not affected and the primary interaction is seen to be electrostatic in nature due to the polar functional groups present on the protein and GO sheet surface. For the first time we have shown that the interaction between ubiquitin and GO is dynamic in nature with fast and reversible adsorption/desorption of protein from the surface of GO. This insight will help in understanding the mechanistic aspects of interaction of GO with cellular proteins and will help in designing appropriate functionalized graphene oxide for its biological application. Chapter 5: Section A: Interaction of an intrinsically disordered protein (L-HIGFBP2) with graphene oxide The interaction between intrinsically disordered linker domain of human insulin-like growth factor binding protein-2 (L-hIGFBP2) with GO was studied using NMR spectroscopy and other techniques such as isothermal titration calorimetry (ITC), dynamic light scattering (DLS), zeta-potential measurements. The study revealed that the disordered protein L-hIGFBP2 interacts with GO through electrostatic interaction and undergoes a dynamic and reversible association-dissociation in a fast exchange regime. The conformation of the protein is not affected. Section B: Stability of an Intrinsically disordered protein through weak interaction with Silver nanoparticles Using NMR spectroscopy and other techniques we probed the mechanism of L-hIGFBP2–AgNP interactions which render the IDP stable. The study reveals a mechanism which involves a relatively fast and reversible association–dissociation of L-hIGFBP2 (dynamic exchange) from the surface of AgNP. The AgNP–L-hIGFBP2 complex remains stable for more than a month. The techniques employed in addition to NMR include UV-Visible spectroscopy, dynamic light scattering (DLS), zeta potential measurements and transmission electron microscopy (TEM) to probe the protein-AgNP interaction here in this section.

Hydrogen bonding properties of water molecules, which are confined in microcavities of biological interfaces, are significantly different from those of bulk water and drive most of the complex biological processes. While NMR, X-ray and UV–vis-IR spectroscopic techniques have been found inadequate for describing the dynamics of the thick (20–40 Å) sheath of hydration layer around biomolecules, recently developed THz spectroscopy has emerged as a powerful technique to directly probe the collective dynamics of hydrogen bonds in the hydration layer, which control all important functions of the biomolecules in life. Both laser and accelerator-based THz sources are intense enough to penetrate up to about 100 μm thick water samples, which makes THz transmission and/or dielectric relaxation measurements possible in aqueous solutions. These measurements provide valuable information about the rattling and rotational motions of hydrated ions, making, breaking and rearrangement of hydrogen bonds in hydration layer as well as hydrophilic and hydrophobic interactions between biomolecule and water. THz spectroscopy has also been successfully applied to study the effect of modulation of the physical conditions, like temperature, pH, concentration of proteins and chemical additives, on the structure and dynamics of hydration layer. THz spectroscopy has also been applied to study the processes of denaturation, unfolding and aggregation of biomolecules.

The biological functions of proteins all depend on their highly specific interactions with other molecules, and the understanding of the molecular basis of the specificity of these interactions is an important part of the effort to understand protein structure-function relationships. NMR spectroscopy can provide information on many different aspects of protein-ligand interactions, ranging from the determination of the complete structure of a protein-ligand complex to focussing on selected features of the interactions between the ligand and protein by using “reporter groups” on the ligand or the protein. It has two particular advantages: the ability to study the complex in solution, and the ability to provide not only structural, but also dynamic, kinetic and thermodynamic information on ligand binding. Early analyses of ligand binding (Jardetzky and Roberts, 1981) focused on measurements of relaxation times, chemical shifts and coupling constants, which gave relatively limited, although valuable, structural information. More recently, it has become possible to obtain much more detailed information, due to the extensive use of nuclear Overhauser effect measurements and isotope-labeled proteins and ligands; a number of reviews of this area are available (Feeney and Birdsall, 1993; Lian et al, 1994; Wand and Short, 1994; Petros and Fesik, 1994; Wemmer and Williams, 1994). In this article, we describe some recent work from our laboratory which illustrates the use of NMR spectroscopy to obtain structural and mechanistic information on relatively large enzyme-substrate and proteinprotein complexes. A number of species of pathogenic bacteria, notably Streptococci and Staphylococci, have proteins on their surface that bind immurioglobulins (reviewed in Boyle (1990)). Protein A from S. aureus and protein G from species of Streptococci are widely used as imrnunological tools and are the most extensively studied of these antibody-binding proteins. A detailed understanding of the binding mechanisms of these proteins is important, not only for providing us with the structural basis for their functions, but also as a contribution toward understanding the general rules of protein-protein interactions.

Fluorescence spectrometry is the most extensively used optical spectroscopic method in analytical measurement and scientific investigation. During the past five years more than 60000 scientific articles have been published in which fluorescence spectroscopy has been used. The large number of applications ranges from the analytical determination of trace metals in the environment to pH measurements in whole cells under physiological conditions. In the scientific research laboratory, fluorescence spectroscopy is being used or applied to study the fundamental physical processes of molecules; structure-function relationships and interactions of biomolecules such as proteins and nucleic acids; structures and activity within whole cells using such instrumentation as confocal microscopy; and DNA sequencing in genomic characterization. In analytical applications the use of fluorescence is dominant in clinical laboratories where fluorescence immunoassays have largely replaced radioimmunoassay techniques. There are two main reasons for this extensive use of fluorescence spectroscopy. Foremost is the high level of sensitivity and wide dynamic range that can be achieved. There are a large number of laboratories that have reported single molecule detection. Secondly, the instrumentation required is convenient and for most purposes can be purchased at a modest cost. While improvements and advances continue to be reported fluorescence instrumentation has reached a high level of maturity. A review of the physical principles of the fluorescence phenomenon permits one to understand the origins of the information content that fluorescence measurements can provide. A molecule absorbs electromagnetic radiation through a quantum mechanical process where the molecule is transformed from a ‘ground’ state to an ‘excited’ state. The energy of the absorbed photon of light corresponds to the energy difference between these two states. In the case of light in the ultraviolet and visible spectral range of 200 nm to 800 nm that corresponds to energies of 143 to 35.8 kcal mol-1. The absorption of light results in an electronic transition in the atom or molecule. In atoms this involves the promotion of an electron from an outer shell orbital to an empty orbital of higher energy.

Pressure-tuning vibrational spectroscopy was first introduced to the study of structural and dynamic properties in biological systems from our laboratory about one decade ago. One of our efforts has been the search for spectral features and their pressure dependencies related to the structural and dynamic properties in biological systems. Pressure-induced correlation field splitting of the vibrational modes of methylene chains is one of the parameters that has been applied to monitor various structural and dynamic properties of a wide range of aqueous lipid bilayers and biomembranes in our laboratory. Correlation field splitting of the vibrational modes of the methylene chains in lipid bilayers is the result of vibrational coupling interactions among the ordered methylene chains with different site symmetry in the two-dimensional matrix. However, the basic theory and the characteristics of these interchain interactions in lipid bilayers still needed to be established. It was unknown whether the interchain interactions that result in the correlation field splitting take place within each lipid molecule or between neighboring molecules in the lamellar bilayers. The relative contributions of 'intramolecular and intermolecular interchain interactions to the correlation field splitting, and the effects of the long-range interchain interactions and interdigitation on the correlation field splitting, were also unknown. These problems have been resolved recently and are addressed in this chapter. Our laboratory has pioneered the study of structural and dynamic properties of biological systems by means of pressure-tuning vibrational spectroscopy (Wong et al., 1982). It is now well recognized that this spectroscopic technique is one of the most powerful physical methods for the study of biological and biomedical phenomena at the molecular level with enhanced resolution (Wong, 1984, 1987a, 1987b, 1987c, 1993). The biological systems we have studied by this method include not only various aqueous biomolecular assemblies but also whole cells and intact biological tissues (Rigas et al, 1990; Wong, 1984, 1987b, 1987c, 1993; Wong et al., 1991a, 1991b, 1993). We have found that the pressure-induced changes in many spectral features and parameters in both FTIR and Raman spectra of biological systems result from modifications in structure and dynamics at the molecular level.