Download or read book Development of Solid state NMR Spectroscopy for Membrane Proteins and the Application to Vpu from HIV 1 written by Eugene Chun-Chin Lin and published by . This book was released on 2015 with total page 148 pages. Available in PDF, EPUB and Kindle. Book excerpt: Solid-state nuclear magnetic resonance (NMR) spectroscopy is a robust method to solve the structures of membrane proteins in their native bilayer environments. Two systems have been well established: oriented samples (OS) are referred to proteins incorporated in the magnetic aligned bicelles, and rotationally aligned (RA) samples are referred to proteins undergoing fast rotational diffusion in proteoliposomes. 1H-15N and 1H-13C dipolar couplings are measured as the angular restraints for the structure calculations. New separated-local-field experiment, Dual-PISEMO, is developed for OS solid-state NMR to measure 1H-13C dipolar couplings of the uniformly 13C labeled proteins, which provide both backbone and side-chain conformations. In order to obtain the full assignments and restraints in OS solid-state NMR, 1H-irradiations with mismatched conditions are implemented to improve the crucial step: the magnetization transfer between 15N and 13C. Non-uniform sampling (NUS) is an alternative approach to further the experimental time of high-dimensional solid-state NMR experiments. Compressed sensing (CS) is able to reconstruct the spectra with 25 ~ 33% of data with the optimized sampling scheme, and covariance spectroscopy provides promising results with alternative States sampling scheme, which requires 50% of data or less. Viral protein "u" (Vpu) from HIV-1 is a type I membrane protein consisting of a transmembrane domain and a cytoplasmic domain, which are associated with different biological activities to enhance viral infectivity. It is essential to determine the three-dimensional structure of Vpu in order to understand its mechanisms in the molecular level and to develop new classes of anti-viral drugs. Magic-angle spinning (MAS) experiments providing high-sensitivity and high- resolution spectra, are implemented to study Vpu incorporated into proteoliposomes. 15N, 13C chemical shift and 1H-15N, 1H-13C dipolar couplings are measured and converted to equivalent structural restraints.

Download or read book NMR Structural Studies of the Integral Membrane Protein Vpu from Hiv 1 written by Che Ma and published by . This book was released on 2000 with total page 88 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Advances in Biological Solid State NMR written by Frances Separovic and published by Royal Society of Chemistry. This book was released on 2014 with total page 632 pages. Available in PDF, EPUB and Kindle. Book excerpt: Advances in Biological NMR brings the reader up to date with chapters from international leaders of this growing field, covering the most recent developments in the methodology and applications of solid state NMR to studies of membrane interactions and molecular motions

Download or read book Development of Solid state NMR Spectroscopy for Membrane Proteins written by Jiaozhi George Lu and published by . This book was released on 2014 with total page 220 pages. Available in PDF, EPUB and Kindle. Book excerpt: Atomic-resolution membrane protein structures can be determined by solid-state Nuclear Magnetic Resonance (NMR) spectroscopy, and the unique advantage of the approach is that membrane proteins reside in near-native lipid bilayer environment at physiological pH and temperature, which minimizes the potential distortions of the protein structure caused by the environment. Here, the full-length mercury transporter protein, MerF, is the focus of the structural studies, and the protein is an essential part of the bacterial mercury detoxification system that has been exploited as a potential engineering target for mercury bioremediation strategies. The backbone structures of the full-length MerF are determined in two environments, (i) magnetically aligned bicelles by oriented-sample (OS) solid-state NMR and (ii) proteoliposome by rotationally aligned (RA) solid-state NMR; and notably, both environments provide the planar lipid bilayer environment for the protein. The structural study of MerF in aligned bicelle has initially been challenging for the OS solid-state NMR, and consequently, methods have been developed to tackle the two major obstacles, the spectral resolution and resonance assignments. New pulse sequence, MSHOT-Pi4/Pi, has demonstrated a reduction of the 1H resonance line width by more than a factor of two, a significant improvement in spectral resolution. New resonance assignment method, Dipolar Coupling Correlated Isotropic Chemical Shift (DCCICS) Analysis, has been developed that is able to transfer resonance assignment from isotropic NMR methods to OS solid-state NMR spectra. The combined usage of several resonance assignment strategies and special tactics, such as applying DCCICS to the new high-resolution proton-evolved local field experiments for terminal and loop residues, has resulted in the complete assignment of all backbone immobile residues of the full-length MerF protein in magnetically aligned bicelle. Meanwhile, RA solid-state NMR is developed in the lab as a new method that combines the strength of magic-angle-spinning (MAS) solid-state NMR in obtaining resonance assignment and the concept of molecular alignment from OS solid-state NMR in obtaining angular restraints. In applying to the structural study of MerF, the method is further incorporated with multi-contact cross polarization and sequential backbone "walk" with three three-dimensional experiments, and the first structure of full-length MerF is determined with the method. In comparison to the previously determined structure of the truncated MerF (MerFt), the full-length structure reveals that the protein truncation has caused large conformational rearrangement at a place more than ten residues away from the truncation site, which serves as an example to demonstrate the importance of studying the full-length unmodified proteins by structural biologists. Additionally, the structure reveals that both mercury-binding sites are located at the intracellular side of the membrane, hinting at the observation of a conformation that allows intramolecular transfer of mercury ions. Subsequently after the complete assignment of MerF in OS solid-state NMR, the MerF structure determined by RA solid-state NMR is further improved by incorporating additional angular restraints from OS solid-state NMR and by the new treatment of dihedral restraints derived from the experimental study of C-terminal dynamics. Lastly, as a side project, the theoretical foundation of MSHOT-Pi4 pulse sequence is further explored. The observation that the pulse sequence selectively improves the resolution of membrane protein samples but not of standard single crystal sample has been analytically generalized as the principle of "motion-adapted" pulse sequence, where it is found that the interference between sample's spatial rotational motion and the radio-frequency pulse rotation in the quantum spin space is the cause of the selectivity. As a related endeavor, the mechanisms of dilute spin exchange and the magic-angle 1H spin-lock pulses have been analyzed theoretically and demonstrated in standard and biological samples. Mixed-order proton-relay mechanism is proposed to be the main contributor to dilute spin exchange in stationary aligned sample, and once more, the difference of pulse performance between standard and biological samples is observed that may be a consequence of several causes including sample motion. In conclusion, the development of various methods in OS and RA solid-state NMR are likely to find their usage in future structural studies of membrane proteins; the theoretical principle of motion-adapted property opens up new avenue to develop pulse sequences for membrane protein samples; and the atomic-resolution backbone structures of MerF contribute information for structural biologist and for the mechanistic study of mercury transportation.

Download or read book Solid state NMR written by Frances Separovic and published by . This book was released on 2020 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: The purpose of this book is to describe the methodology and applications of solid-state NMR spectroscopy to studies of membrane proteins, membrane-active peptides and model biological membranes. As well as structural studies, this book contains coverage of membrane interactions and molecular motions. Advances in biological solid-state NMR are very pertinent with high-field developments seeing applications in biological membranes and whole cells. Experts who are leaders in the development and application of biological solid-state NMR are chapter contributors. Part of Biophysical Society-IOP series.

Download or read book Perspectives on Solid State NMR in Biology written by S.R. Kiihne and published by Springer Science & Business Media. This book was released on 2001-07-31 with total page 256 pages. Available in PDF, EPUB and Kindle. Book excerpt: Solid state NMR is rapidly emerging as a universally applicable method for the characterization of ordered structures that cannot be studied with solution methods or diffraction techniques. This proceedings -; from a recent international workshop - captures an image of the latest developments and future directions for solid state NMR in biological research, particularly on membrane proteins. Detailed information on how hormones or drugs bind to their membrane receptor targets is needed, e.g. for rational drug design. Higher fields are bringing clear improvements, and the power of solid state NMR techniques for studying amorphous and membrane associated peptides, proteins and complexes is shown by examples of applications at ultra-high fields. Progress in protein expression, experimental design and data analysis are also presented by leaders in these research areas.

Download or read book Solid state NMR Spectroscopy of HIV 1 Vpu and Human CD4 in Lipid Bilayers written by Hoa Quynh Do and published by . This book was released on 2012 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Solid State NMR Spectroscopy for Biopolymers written by Hazime Saitô and published by Springer. This book was released on 2009-09-03 with total page 455 pages. Available in PDF, EPUB and Kindle. Book excerpt: ‘‘Biopolymers’’ are polymeric materials of biological origin, including globular, membrane, and fibrous proteins, polypeptides, nucleic acids, po- saccharides, lipids, etc. and their assembly, although preference to respe- ive subjects may be different among readers who are more interested in their biological significance or industrial and/or medical applications. Nevert- less, characterizing or revealing their secondary structure and dynamics may be an equally very important and useful issue for both kinds of readers. Special interest in revealing the 3D structure of globular proteins, nucleic acids, and peptides was aroused in relation to the currently active Structural Biology. X-ray crystallography and multidimensional solution NMR sp- troscopy have proved to be the standard and indispensable means for this purpose. There remain, however, several limitations to this end, if one intends to expand its scope further. This is because these approaches are not always straightforward to characterize fibrous or membrane proteins owing to extreme difficulty in crystallization in the former, and insufficient spectral resolution due to sparing solubility or increased effective molecular mass in the presence of surrounding lipid bilayers in the latter.

Download or read book Protein NMR written by Lawrence Berliner and published by Springer. This book was released on 2015-08-24 with total page 193 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book covers new techniques in protein NMR, from basic principles to state-of-the-art research. It covers a spectrum of topics ranging from a “toolbox” for how sequence-specific resonance assignments can be obtained using a suite of 2D and 3D NMR experiments and tips on how overlap problems can be overcome. Further topics include the novel applications of Overhauser dynamic nuclear polarization methods (DNP), assessing protein structure, and aspects of solid-state NMR of macroscopically aligned membrane proteins. This book is an ideal resource for students and researchers in the fields of biochemistry, chemistry, and pharmacology and NMR physics. Comprehensive and intuitively structured, this book examines protein NMR and new novel applications that include the latest technological advances. This book also has the features of: • A selection of various applications and cutting-edge advances, such as novel applications of Overhauser dynamic nuclear polarization methods (DNP) and a suite of 2D and 3D NMR experiments and tips on how overlap problems can be overcome • A pedagogical approach to the methodology • Engaging the reader and student with a clear, yet critical presentation of the applications

Download or read book Biomedical Technology Resources written by and published by . This book was released on 1996 with total page 130 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Characterization of Structure and Dynamics of Membrane Proteins from Solid state NMR written by Byungsu Kwon and published by . This book was released on 2018 with total page 144 pages. Available in PDF, EPUB and Kindle. Book excerpt: Solid-state nuclear magnetic resonance (ssNMR) spectroscopy is an essential tool for elucidating the structure, dynamics, and function of biomolecules. ssNMR is capable of studying membrane proteins in near-native lipid bilayers and is thus preferred over other biophysical techniques for characterizing the structure and dynamics of membrane proteins. This thesis primarily focuses on the study of the following membrane proteins: 1) the N-terminal ectodomain and C-terminal cytoplasmic domain of influenza A virus M2 and 2) HIV-1 glycoprotein gp4l membrane-proximal external region and transmembrane domain (MPER-TMD) in a near native membrane environment. The cytoplasmic domain of M2 is necessary for membrane scission and virus shedding. The M2(22-71) construct shows random-coil chemical shifts, large motional amplitudes, and a membrane surface-bound location with close proximity to water, indicating the post-amphipathic helix (AH) cytoplasmic domain is a dynamic random coil near the membrane surface. The influenza M2 ectodomain contains highly conserved epitopes but its structure is largely unknown. The M2(1-49) construct containing both the ectodomain and transmembrane domain exhibits an entirely unstructured ectodomain with a motional gradient in which the motion is slower for residues near the TM domain, which attributed to the formation of a tighter helical bundle in the presence of drug that should cause the more tightened C-terminal ectodomain, thereby slowing its local motions. HIV-1 virus gp4l is directly involved in virus-cell membrane fusion. However, the structural topologies of the gp4l MPER-TMD are still controversial and the biologically-relevant intrinsic conformational state of MPER has not yet been determined. In order to obtain near native structural information of gp4l, we have studied gp41 (665-704) and found a primarily a-helical conformation, membrane-anchored trimeric TMD and water-exposed membrane surface-bound MPER. Intra- and intermolecular distances measured using 19C-19F REDOR and 19F-19F CODEX revealed that MPER-TMD has a significant kink between MPER and TMD, which has aided a deeper understanding of the HIV virus entry mechanism and the design of vaccines.

Download or read book Protein NMR Techniques written by A. Kristina Downing and published by Springer Science & Business Media. This book was released on 2008-02-03 with total page 494 pages. Available in PDF, EPUB and Kindle. Book excerpt: When I was asked to edit the second edition of Protein NMR Techniques, my first thought was that the time was ripe for a new edition. The past several years have seen a surge in the development of novel methods that are truly revolutionizing our ability to characterize biological macromolecules in terms of speed, accuracy, and size limitations. I was particularly excited at the prospect of making these techniques accessible to all NMR labs and for the opportunity to ask the experts to divulge their hints and tips and to write, practically, about the methods. I commissioned 19 chapters with wide scope for Protein NMR Techniques, and the volume has been organized with numerous themes in mind. Chapters 1 and 2 deal with recombinant protein expression using two organisms, E. coli and P. pastoris, that can produce high yields of isotopically labeled protein at a reasonable cost. Staying with the idea of isotopic labeling, Chapter 3 describes methods for perdeuteration and site-specific protonation and is the first of several chapters in the book that is relevant to studies of higher molecular weight systems. A different, but equally powerful, method that uses molecular biology to “edit” the spectrum of a large molecule using segmental labeling is presented in Chapter 4. Having successfully produced a high molecular weight target for study, the next logical step is data acquisition. Hence, the final chapter on this theme, Chapter 5, describes TROSY methods for stru- ural studies.

Download or read book Solution and Solid state NMR of Membrane bound Proteins and Peptides written by Alessandro Mascioni and published by . This book was released on 2003 with total page 670 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Solid state NMR Studies of Structure and Dynamics of HIV 1 Capsid CA Protein Assemblies written by Yun Han and published by . This book was released on 2014 with total page 211 pages. Available in PDF, EPUB and Kindle. Book excerpt: There are around 34 million people in the world living with HIV-1, which is a causative agent of acquired immunodeficiency syndrome (AIDS). AIDS has become the ninth leading cause for death of people ages 25-34 in the US. Even though highly active antiretroviral therapy (HAART) showed effectiveness in suppressing the virus replication and significantly prolonged the patients lives, AIDS still remains an uncured disease. To develop new therapies, atomic-level understanding of the mechanism of HIV-1 lifecycle, including the structures of the various protein assemblies, is needed. The Gag polyprotein and its component capsid (CA) protein are essential constituents of the HIV-1 life cycle, and have recently attracted attention as targets for drug development. However, the atomic resolution structure and the dynamics of Gag and CA protein assemblies and their complexes with small-molecule inhibitors are not available because these assemblies are not amenable for characterization by traditional structural biology methods, X-ray diffraction and solution NMR spectroscopy. Solid-state NMR spectroscopy has the unique capability of providing atomic-level structural and dynamics information in large protein assemblies. The focus of this dissertation is establishing solid-state NMR spectroscopy as an atomic-level probe of structure and dynamics in HIV-1 protein assemblies. This effort required first establishing sample conditions for formation of HIV-1 protein assemblies that give rise to high-resolution solid-state NMR spectra for subsequent structural studies. In my Ph. D. work, I have optimized protocols to prepare homogeneous HIV-1 CA protein assemblies in vitro, developed confocal imaging method to characterize the morphologies of the resulting assemblies. With the suitable samples in hand, I have acquired solid-state NMR spectra on assemblies of CA protein and the maturation intermediate, CA-SP1, for structural analysis. Using these solid-state NMR data, I and my colleagues have obtained novel insights into the following aspects of HIV-1 structural biology: i) conformation of CA protein in conical assemblies; ii) the role of conformational dynamics of the hinge region in the structural polymorphism of CA in conical assemblies; iii) conformation of spacer peptide SP1 in tubular CA-SP1 assemblies; iv) conformation and dynamics of CA in tubular assemblies. In this thesis, I will first discuss the preparation and characterization of HIV-1 CA and CA-SP1 protein assemblies (Chapter 2 and 3). I will then describe the resonance assignments and secondary structure analysis of conical assemblies of HIV-1 CA protein (Chapter 4). Next, in Chapter 5, I will present the dynamics studies of conical assemblies of HIV-1 CA protein assemblies. In Chapter 6, I will discuss the resonance assignments and conformational analysis of tubular assemblies of HIV-1 CA and CA-SP1 proteins. The long-term goal of this research is developing comprehensive understanding of the mechanism of capsid assembly and disassembly, HIV-1 maturation, through the structural and dynamics analysis of the CA and Gag assemblies. The work discussed here represents the first step toward this goal and lays out the methodological and intellectual foundations enabling the solid-state NMR analysis of HIV-1 protein assemblies.

Download or read book NMR of Proteins and Small Biomolecules written by Guang Zhu and published by Springer Science & Business Media. This book was released on 2012-03-28 with total page 255 pages. Available in PDF, EPUB and Kindle. Book excerpt: Application of NMR and Molecular Docking in Structure-Based Drug Discovery, by Jaime L. Stark and Robert Powers NMR as a Unique Tool in Assessment and Complex Determination of Weak Protein-Protein Interactions, by Olga Vinogradova and Jun Qin The Use of Residual Dipolar Coupling in Studying Proteins by NMR, by Kang Chen und Nico Tjandra NMR Studies of Metalloproteins, by Hongyan Li and Hongzhe Sun Recent Developments in 15N NMR Relaxation Studies that Probe Protein Backbone Dynamics, by Rieko Ishima Contemporary Methods in Structure Determination of Membrane Proteins by Solution NMR, by Tabussom Qureshi and Natalie K. Goto Protein Structure Determination by Solid-State NMR, by Xin Zhao Dynamic Nuclear Polarization: New Methodology and Applications, by Kong Hung Sze, Qinglin Wu, Ho Sum Tse and Guang Zhu

Download or read book Development of Solid state NMR Methodologies for Protein Structure Determination Based on Paramagnetic Tagging written by Dwaipayan Mukhopadhyay and published by . This book was released on 2018 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: In recent years solid-state NMR has emerged as a valuable tool for elucidating structure and dynamics of large biomolecular systems, especially suited for application in systems ranging from amyloid fibrils, membrane proteins to large protein ligand complexes inaccessible by more traditional structure determination techniques e.g. X-ray crystallography and solution-state NMR. Essential in determination of a biomolecular structure is the acquisition of numerous high quality interatomic distance restraints. However, conventional dipolar coupling based methods lead to significant challenges for accurate determination of critical long range distances. An alternative method to overcome this limitation is the incorporation of paramagnetic centers in proteins and utilization of the very large electron-nucleus couplings. In this dissertation, we combine state of the art proton-detection techniques with development of novel rigid transition metal binding tags, to improve the quality of long range distance restraints generated by measurement of site-specific paramagnetic relaxation enhancements in solid-state NMR.

Download or read book Mas Solid State Nmr on Biomolecules written by Rasmus Linser and published by Sudwestdeutscher Verlag Fur Hochschulschriften AG. This book was released on 2011 with total page 192 pages. Available in PDF, EPUB and Kindle. Book excerpt: Due to numerous achievements in the last years, solid-state NMR has evolved a potent tool for structural characterization of biomolecules. Not relying on single crystals or solubility, the technique particularly focuses on fibrillar and membrane proteins, which are difficult to characterize otherwise. Despite this enormous potential, numerous difficulties unsolved to date demand for further technical and methodological reworking. Among these, the achievable signal to noise and resolution are the most prominent limits. Whereas a series of limitations is induced by strong dipolar interactions among protons even under fast MAS (Magic Angle Spinning), a (partial) dilution of the proton content by deuterons bears a number of spectroscopic advantages. These are e.g. direct 1H detection with an excellent resolution, accessibility of long-range distance restraints, and proton based dynamics and surface accessibility determination. Besides, relaxation for various coherences is significantly decreased, making solution-NMR like multi-dimensional experiments for peak assignment etc. possible. The features of a dilute protonation explored in this work have lead to 12 peer-reviewed publications.