Use of Beta-lactamase Fusions to Study Membrane Protein Assembly and Protein Export in Escherichia Coli PDF Download
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Author: Yianbiao Zhang
Publisher:
ISBN:
Category :
Languages : en
Pages :
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Book Description
Author: Yianbiao Zhang
Publisher:
ISBN:
Category :
Languages : en
Pages :
Get Book Here
Book Description
Author:
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Book Description
A genetic system for directly expressing proteins in E. coli and assessing their ability to insert into the bacterial cytoplasmic membrane was developed, consisting of a direct expression vector, and a mature P-lactamase cassette plasmid for generating translational fusions of P-lactamase to the bacterially expressed proteins. This system was used to analyse the fate of three different eukaryotic plasma membrane proteins in E. coli. The ampicillin resistance/sensitivity phenotypes of the P-lactamase fusion proteins revealed that the P-subunit of the sheep-kidney Na,K-ATPase, a class II membrane protein, inserts into the cytoplasmic membrane of E. coli, with the same topology as in its native membrane. The Alzheimer's disease amyloid A4 protein precursor, a class I membrane protein, was also found to insert into the bacterial cytoplasmic membrane, and, although preliminary, the results were compatible with its correct topological insertion into the bacterial cytoplasmic membrane. In contrast, the P2-adrenergic receptor, a complex membrane protein that has an extracellular amino-terminal domain and seven transmembrane segments, was found to misinsert into the E. coli cytoplasmic membrane. A plasmid vector for cloning sequences able to function as cytoplasmic membrane translocators of P-lactamase in E. coli was also developed. Short DNA fragments of essentially random sequence were cloned, and recombinants with functional translocators were directly selected. The spectrum of random sequences able to function as bacterial cytoplasmic membrane translocators suggests that the hydrophobicity requirements for translocation across the bacterial cytoplasmic membrane and yeast endoplasmic reticulum membrane may be more similar than previously assumed.In the above studies new and widely applicable uses of P-lactamase fusion technology have been described. As applied in this thesis they highlight similarities and differences between the mechanisms by which proteins assemble into, or are translocated across, the bacterial cytoplasmic membrane and the eukaryotic endoplasmic reticulum membrane.
Author: R.E. Dalbey
Publisher: Elsevier
ISBN: 0080526098
Category : Science
Languages : en
Pages : 297
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Book Description
The incentive for putting together Volume 4 of this series was to review the wealth of new information that has become available in prokaryotic organisms in protein export and membrane biogenesis. Just in the last several years, protein translocation has now been efficiently reconstituted using defined components and the mechanism by which proteins are moved across membrane bilayers is now being examined at a higher resolution. In addition, because of a new technical breakthrough using osmolytes, it is now possible to reconstitute a number of channel proteins, ATPase, receptors, and transporters. In many cases, it is possible to successfully predict the membrane topology of these types of proteins using both "hydrophobicity analysis" and the "positive inside" rule. In this volume, two chapters focus on protein translocation across membranes (Biochemical Analyses of Components Comprising the Protein Translocation Machinery of E. Coli; Protein Translocation Genetics), while several others on how proteins assemble into the ineer membrane of E. Coli (Membrane Protein Assembly; Membrane Insertion of Small Proteins: Evolutionary and Functional Aspects; Pigment-Protein Complex Assembly in Rhodobacter sphaeroides and Rhodobacter Capsulatus). Other sections review recent progress on transporters (Identification and Reconstitution of Anion Exchange Mechanisms in Bacteria; Helic Packing in the C-Terminal Half of Lactose Permease) and signal transduction (Mechanism of Transmembrane Signaling in Osmoregulation) as well as the assembly of prints into the outer membrane (Export and Assembly of Outer Membrane Proteins in E. coli). Although the emphasis of the book is on proteins, the role of phospholipids in controlling various cell surface processes is reviewed (Role of Phospholipids in coli Cell Function). I should point out the reason for the rapid progress in bacteria research is because of the possibility to apply biochemistry and genetics in this organism.
Author: Jos A.F. Op den Kamp
Publisher: Springer Science & Business Media
ISBN: 3642731848
Category : Science
Languages : en
Pages : 474
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Book Description
Many individual aspects of the dynamics and assembly of biological membranes have been studied in great detail. Cell biological approaches, advanced genetics, biophysics and biochemistry have greatly contributed to an increase in our knowledge in this field.lt is obvious however, that the three major membrane constituents - lipids, proteins and carbohydrates- are studied, in most cases separately and that a coherent overview of the various aspects of membrane biogenesis is not readily available. The NATO Advanced Study Institute on "New Perspectives in the Dynamics of Assembly of Biomembranes" intended to provide such an overview: it was set up to teach students and specialists the achievements obtained in the various research areas and to try and integrate the numerous aspects of membrane assembly into a coherent framework. The articles in here reflect this. Statting with detailed contributions on phospholipid structure, dynamics, organization and biogenesis, an up to date overview of the basic, lipidic backbone of biomembranes is given. Extensive progress is made in the research on membrane protein biosynthesis. In particular the post- and co-translational modification processes of proteins, the mechanisms of protein translocation and the sorting mechanisms which are necessary to direct proteins to their final, intra - or extracellular destination have been characterized in detail. Modern genetic approaches were indispensable in this research area: gene cloning, hybrid protein construction, site directed mutagenesis and sequencing techniques elucidated many functional aspects of specific nucleic acid and amino acid sequences.
Author: Stephen H White
Publisher: Springer
ISBN: 1461475155
Category : Science
Languages : en
Pages : 403
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Book Description
Studies of receptors, ion channels, and other membrane proteins require a solid understanding of the structural principles of these important biomolecules. Membrane protein structure is, however, a very challenging field. The structures of only three types of transmembrane proteins have been determined to moderate or high resolution during the last two decades, a period during which the amino acid sequences of hundreds, if not thousands, of membrane proteins have been reported. As a result, the creation of structural models to serve as guides for studies of receptors, channels, and other membrane proteins has become crucially important. This book has been assembled in order to share the experiences and findings of expert researchers in protein structure and structure-prediction methods as well as membrane biophysics and lipid physical chemistry, whose work establishes the basis for the development of suitable model structures. The reviews presented here emphasize fundamental ideas and provide an entry to the diverse and complex literature. The four major sections deal with the general nature of the membrane protein structure problem, biochemical and molecular biological approaches to protein topology, direct structural methods, and model and physicochemical approaches. The work will be of interest to physiologists, cellular and molecular biologists, biophysicists, and biochemists working on the function of membrane proteins such as receptors, ion channels, and transporters, as well as senior graduate students and independent investigators.
Author: Gunnar von Heijne
Publisher: R. G. Landes
ISBN:
Category : Science
Languages : en
Pages : 300
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Book Description
Author: Henry C. Wu
Publisher: Springer Science & Business Media
ISBN: 3642712517
Category : Medical
Languages : en
Pages : 211
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Book Description
The last decade has witnessed rapid progress in our under standing of the mechanisms of protein export and secretion in both prokaryotic and eukaryotic cells. Studies of protein secretion across the membranes of the rough endoplasmic reticulum have led to the formulation of the now-classic signal hypothesis, which has stimulated many discussions and new ideas, and the identification of the signal recogni tion particle as an organelle in the initiation of the export process. However, more recent work pertaining to intrage nic information related to targeting specific proteins for either secretion or membrane localization, the energetics of protein secretion, the timing of synthesis versus the initia tion of export, structural requirements for the processing of precursor proteins, and the identification of the proces sing enzymes (signal peptidases), has been the result of a combined biochemical and genetic approach to the study of protein localization in bacteria. While reviews on the biochemistry and genetics of pro tein secretion have appeared frequently in recent years, this book attempts to summarize the current status and the future perspectives of this rapidly moving field in a single volume. Topics covered in this book include the genetics of protein secretion in E. coli, biochemical analysis of pro tein export in vitro, signal peptidases, excretion of colicins and hemolysin in E. coli, protein secretion in Bacillus, and protein secretion cloning vectors.
Author:
Publisher:
ISBN:
Category : Dissertations, Academic
Languages : en
Pages : 498
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Book Description
Theses on any subject submitted by the academic libraries in the UK and Ireland.
Author: Joseph Bentz
Publisher: CRC Press
ISBN: 9780849356063
Category : Science
Languages : en
Pages : 548
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Book Description
Viral Fusion Mechanisms presents the first comprehensive review on this exciting topic. The book focuses on molecular mechanisms rather than phenomonology and examines a wide range of viruses, including influenza, HIV, Sendai, SFV, Vaccinia, VSV, and RSV. Recent theoretical work on dissecting protein-mediated membrane fusion is discussed, and the most promising new technologies for elucidating mechanisms are highlighted. Viral Fusion Mechanisms is an essential reference for biophysicists, cell biologists, colloid chemists, immunologists, microbiologists, molecular biologists, and virologists.
Author: Anne Skaja Robinson
Publisher: John Wiley & Sons
ISBN: 3527634533
Category : Science
Languages : en
Pages : 631
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Book Description
Designed as a research-level guide to current strategies and methods of membrane protein production on the small to intermediate scale, this practice-oriented book provides detailed, step-by-step laboratory protocols as well as an explanation of the principles behind each method, together with a discussion of its relative advantages and disadvantages. Following an introductory section on current challenges in membrane protein production, the book goes on to look at expression systems, emerging methods and approaches, and protein specific considerations. Case studies illustrate how to select or sample the optimal production system for any desired membrane protein, saving both time and money on the laboratory as well as the technical production scale. Unique in its coverage of "difficult" proteins with large membrane-embedded domains, proteins from extremophiles, peripheral membrane proteins, and protein fragments.
