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Author: Suhani Nagpal
Publisher:
ISBN:
Category :
Languages : en
Pages : 382
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Book Description
In order to execute their biological activities, most proteins fold into their unique, three-dimensional structure. The discovery of intrinsically disordered proteins (IDPs) about two decades ago, which are now widely found in eukaryotes, has since challenged the structure-function paradigm. IDPs, which in isolation exist as broad, non-random, conformational ensembles of interconverting states, are centrally involved in many biological processes. The key to their functioning is the ability to fold when bound to ligand partner(s), thus operating as morphing proteins. Despite booming interest in morphing behavior, investigating their structural transitions and mechanism remains extremely difficult because of their distinct characteristics. Previously, we observed a close connection between intrinsically partially disordered proteins (IPDPS) and gradual (un)folding transitions of downhill folders, leading to the hypothesis that many IPDPs work as a conformational rheostat. The scope of this dissertation is to investigate the biological and technological implications of gradual conformational transitions. We first demonstrate the design principles of protein-based scaffolds by utilizing gradual (un)folding coupled to binding for developing rheostatic conformational transducers using computational modeling and experiments. Our engineered transducers showcase>6 orders of magnitude change in analyte concentration (broadband sensitivity) and have practical advantages over extant ones, which conventionally operate as conformational switches. Next, inspired by the LEGO toy, we devised a novel modular approach to dissect the folding cooperativity and the energetic contributions of native interactions in defining the conformational ensemble and binding properties of IPDPs. Using an integrated strategy of computation and experiments, we perform an ensemble-based conformational analysis and find that the approach provides an exciting new tool for analyzing morphing transitions that should generally apply to any IPDP, thereby addressing a fundamental gap in the field. One particularly interesting IPDP is NCBD that binds to multiple structurally diverse ligand partners and recruits the basal transcription machinery. We then explore the concept of NCBD functioning as a conformational rheostat, which allows its promiscuous binding. Finally, using extensive all-atom Molecular Dynamics simulations of NCBD and its biological partners in their free and bound forms, we decipher the hidden conformational biases in the dynamics of the heterogeneous ensemble of NCBD, undergoing gradual morphing transitions hinting at a working conformational rheostat in transcription.
Author: Suhani Nagpal
Publisher:
ISBN:
Category :
Languages : en
Pages : 382
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Book Description
In order to execute their biological activities, most proteins fold into their unique, three-dimensional structure. The discovery of intrinsically disordered proteins (IDPs) about two decades ago, which are now widely found in eukaryotes, has since challenged the structure-function paradigm. IDPs, which in isolation exist as broad, non-random, conformational ensembles of interconverting states, are centrally involved in many biological processes. The key to their functioning is the ability to fold when bound to ligand partner(s), thus operating as morphing proteins. Despite booming interest in morphing behavior, investigating their structural transitions and mechanism remains extremely difficult because of their distinct characteristics. Previously, we observed a close connection between intrinsically partially disordered proteins (IPDPS) and gradual (un)folding transitions of downhill folders, leading to the hypothesis that many IPDPs work as a conformational rheostat. The scope of this dissertation is to investigate the biological and technological implications of gradual conformational transitions. We first demonstrate the design principles of protein-based scaffolds by utilizing gradual (un)folding coupled to binding for developing rheostatic conformational transducers using computational modeling and experiments. Our engineered transducers showcase>6 orders of magnitude change in analyte concentration (broadband sensitivity) and have practical advantages over extant ones, which conventionally operate as conformational switches. Next, inspired by the LEGO toy, we devised a novel modular approach to dissect the folding cooperativity and the energetic contributions of native interactions in defining the conformational ensemble and binding properties of IPDPs. Using an integrated strategy of computation and experiments, we perform an ensemble-based conformational analysis and find that the approach provides an exciting new tool for analyzing morphing transitions that should generally apply to any IPDP, thereby addressing a fundamental gap in the field. One particularly interesting IPDP is NCBD that binds to multiple structurally diverse ligand partners and recruits the basal transcription machinery. We then explore the concept of NCBD functioning as a conformational rheostat, which allows its promiscuous binding. Finally, using extensive all-atom Molecular Dynamics simulations of NCBD and its biological partners in their free and bound forms, we decipher the hidden conformational biases in the dynamics of the heterogeneous ensemble of NCBD, undergoing gradual morphing transitions hinting at a working conformational rheostat in transcription.
Author: Thinh Luong
Publisher:
ISBN:
Category :
Languages : en
Pages : 204
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Book Description
Proteins are the work horses of the cell that perform the vast majority of functions essential for life. The mechanism by which proteins fold to their functional native state has been a subject of extensive research for more than 50 years now. Downhill folders are the class of proteins whose folding reaction is heterogeneous, non-cooperative, and happens without encountering a significant free energy barrier, resulting in ultrafast kinetics. The single ensemble of conformations of a global downhill folding protein moves gradually from highly disordered to the unique native structure when thermodynamic parameters that affect the protein's stability are changed (one-state folding). The gradual morphing of a one-state downhill folding protein structure in response to thermodynamic bias is referred to as a conformational rheostat. When such a conformational rheostat is coupled to binding an analyte, it can result in an ultrafast, broad band, and single-molecule analog biosensors. This thesis explores conformational rheostats as the mechanism behind the folding upon binding behavior of intrinsically disordered proteins and as broadband transducers towards engineering high-performance biosensors.The second chapter of this thesis describes a new methodology that we have developed to study the conformational landscape of intrinsically partially disordered proteins (IPDP). This methodology is inspired by the LEGO game, where the sequence of an IPDP is deconstructed into its local structural elements and their possible combinations based on the 3D structure the IPDP acquires upon binding its partners. The local structural elements are hence analogous to LEGO building blocks, and their combinations report on the interactions among them, like the complementary indentations of LEGO pieces. In particular, we chose the IPDP NCBD as model IPDP to develop the proof of concept for the method. Our results showed that even though the NCBD is highly flexible and apparently disordered, there are strong local signals and different sets of long-range transient interactions. These sets of interactions stabilize the overall fold and compete with one another hence resulting in a dynamic ensemble. The methodology developed in Chapter 2 is expected to be extremely useful in characterizing the incipient cooperativity of virtually any IPDP in their unbound form, a capability that is currently unavailable. Chapters 3 and 4 of this thesis deal with the design of a pH biosensor using the downhill folding protein gpW as a scaffold and unfolding coupled to ionization as a transducer. In chapter 3, a methodology for engineering conformational pH transducing into pH insensitive proteins using a histidine grafting approach was developed. The methodology was applied to the protein gpW to demonstrate an engineered, tunable broadband pH transducer based on the conformational rheostat mechanism. Chapter 4 explores general strategies for introducing fluorescence readouts capable of converting the gradual conformational changes of the rheostatic pH transducer into broadband fluorescence-based pH biosensors. Strategies that exploit the Förster Resonance Energy Transfer (FRET) and Photo Induced Electron Transfer (PET) mechanisms were explored as potential means to convert changes in conformation into suitable fluorescence signals were explored and characterized. We discovered that FRET signals using fluorophores in the visible (required for high-sensitivity biosensing) are insensitive to the localized conformational changes associated with conformational rheostats in native-like conditions. In contrast, the very short-range distance dependence of PET (
Author: Derek J. Chadwick
Publisher: John Wiley & Sons
ISBN: 0470514159
Category : Science
Languages : en
Pages : 282
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Book Description
How the amino acid sequence of a protein determines its three-dimensional structure is a major problem in biology and chemistry. Leading experts in the fields of NMR spectroscopy, X-ray crystallography, protein engineering and molecular modeling offer provocative insights into current views on the protein folding problem and various aspects for future progress.
Author: Barry T. Nall
Publisher:
ISBN:
Category : Science
Languages : en
Pages : 236
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Book Description
Author: David Edward Wildes
Publisher:
ISBN:
Category :
Languages : en
Pages : 402
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Book Description
Author: Charis Ghelis
Publisher: Academic Press
ISBN: 0323140920
Category : Science
Languages : en
Pages : 580
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Book Description
Protein Folding aims to collect the most important information in the field of protein folding and probes the main principles that govern formation of the three-dimensional structure of a protein from a nascent polypeptide chain, as well as how the functional properties appear. This text is organized into three sections and consists of 15 chapters. After an introductory chapter where the main problems of protein folding are considered at the cellular level in the context of protein biosynthesis, the discussion turns to the conformation of native globular proteins. Definitions and rules of nomenclature are given, including the structural organization of globular proteins deduced from X-ray crystallographic data. Folding mechanisms are tentatively deduced from the observation of invariants in the architecture of folded proteins. The next chapters focus on the energetics of protein conformation and structure, indicating the principles of thermodynamic stability of the native structure, along with theoretical computation studies of protein folding, structure prediction, and folding simulation. The reader is also introduced to various experimental approaches; the reversibility of the unfolding-folding process; equilibrium and kinetic studies; and detection and characterization of intermediates in protein folding. This text concludes with a chapter dealing with problems specific to oligomeric proteins. This book is intended for research scientists, specialists, biochemists, and students of biochemistry and biology.
Author: Ameed H. Hashmi
Publisher:
ISBN:
Category :
Languages : en
Pages : 314
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Book Description
The research described in this thesis aims at exploiting recent important theoretical and experimental developments in protein folding towards developing advanced biosensors with improved/novel properties. Transforming this basic protein folding knowledge into engineering strategies for designing novel biosensors is an enormous challenge. Here we demonstrate the very first steps in that direction. Biosensors based on proteins that naturally toggle between two states (unfolded/folded) upon specific binding to a target molecule have been successfully used for real-time sensing. These devices behave as conformational switch sensors. Principles on how to engineer this type of conformational transducer onto any protein of interest have also been laid out. Our work takes this state of the art forward to design high-performance conformational rheostat sensors. The rationale is to develop sensors with expanded dynamic range and faster response time by using as conformational transducer the coupling of binding to the analyte and the gradual folding process of fast folding protein modules, and fluorescence as optical readout. As proof of concept, we investigate the pH sensing capabilities of engineered proteins based on two scaffolds: i) an anti-parallel coiled-coil, and ii) a tandem array of the small downhill folding domain BBL. Our results reveal that such pH sensors exhibit a linear response over at least 4 orders of magnitude in proton concentration. We also demonstrate that a pH sensor based on a conformational rheostat transducer can produce analog pH readouts at the single-molecule level together with ultrafast response (
Author: Richard A. Friesner
Publisher: John Wiley & Sons
ISBN: 0471465232
Category : Science
Languages : en
Pages : 544
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Book Description
Since the first attempts to model proteins on a computer began almost thirty years ago, our understanding of protein structure and dynamics has dramatically increased. Spectroscopic measurement techniques continue to improve in resolution and sensitivity, allowing a wealth of information to be obtained with regard to the kinetics of protein folding and unfolding, and complementing the detailed structural picture of the folded state. Concurrently, algorithms, software, and computational hardware have progressed to the point where both structural and kinetic problems may be studied with a fair degree of realism. Despite these advances, many major challenges remain in understanding protein folding at both the conceptual and practical levels. Computational Methods for Protein Folding seeks to illuminate recent advances in computational modeling of protein folding in a way that will be useful to physicists, chemists, and chemical physicists. Covering a broad spectrum of computational methods and practices culled from a variety of research fields, the editors present a full range of models that, together, provide a thorough and current description of all aspects of protein folding. A valuable resource for both students and professionals in the field, the book will be of value both as a cutting-edge overview of existing information and as a catalyst for inspiring new studies. Computational Methods for Protein Folding is the 120th volume in the acclaimed series Advances in Chemical Physics, a compilation of scholarly works dedicated to the dissemination of contemporary advances in chemical physics, edited by Nobel Prize-winner Ilya Prigogine.
Author: Bret A. Shirley
Publisher: Taylor & Francis US
ISBN: 9780896033016
Category : Medical
Languages : en
Pages : 392
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Book Description
In Protein Stability and Folding: Theory and Practice, world-class scientists present in a single volume a comprehensive selection of hands-on recipes for all of the major techniques needed to understand the conformational stability of proteins, as well as their three-dimensional folding. The distinguished contributors provide clear, step-by-step instructions along with many troubleshooting tips, alternative procedures, and informative explanations about why certain steps are necessary. Even highly skilled researchers will find many time-saving methods. Among the techniques discussed are fluorescent, ultraviolet, and infrared spectroscopy; HPLC peptide mapping; differential scanning calorimetry; and hydrogen exchange. Shirley's Protein Stability and Folding: Theory and Practice will ensure a significant difference in the outcome of your experiments, producing the result desired even for beginners.
Author: Kenneth M.Jr. Merz
Publisher: Springer Science & Business Media
ISBN: 1468468316
Category : Science
Languages : en
Pages : 585
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Book Description
A solution to the protein folding problem has eluded researchers for more than 30 years. The stakes are high. Such a solution will make 40,000 more tertiary structures available for immediate study by translating the DNA sequence information in the sequence databases into three-dimensional protein structures. This translation will be indispensable for the analy sis of results from the Human Genome Project, de novo protein design, and many other areas of biotechnological research. Finally, an in-depth study of the rules of protein folding should provide vital clues to the protein fold ing process. The search for these rules is therefore an important objective for theoretical molecular biology. Both experimental and theoretical ap proaches have been used in the search for a solution, with many promising results but no general solution. In recent years, there has been an exponen tial increase in the power of computers. This has triggered an incredible outburst of theoretical approaches to solving the protein folding problem ranging from molecular dynamics-based studies of proteins in solution to the actual prediction of protein structures from first principles. This volume attempts to present a concise overview of these advances. Adrian Roitberg and Ron Elber describe the locally enhanced sam pling/simulated annealing conformational search algorithm (Chapter 1), which is potentially useful for the rapid conformational search of larger molecular systems.
