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Author: Pamela Sears Publisher: ISBN: Category : Languages : en Pages : 5
Book Description
Barnase was successfully displayed on the surface of m13 phage as an N-terminal fusion with coat protein III. The successful display was verified by Western Blotting of purified phage. For selection of a thermostable barnase, the buried residues Ala11, Leu14, Leu20, Ala74, Ile88, Tyr90, and Ile96 were replaced randomly with Phe, Leu, Val, Ile, Ala, Pro, Gly, Ser, or Thr, giving a library of 2x10(exp 6) members. The transformation yield obtained (4x10(exp 6) allows sampling of all possible members. We are currently selecting the phage library on immobilized barstar (the femtomolar inhibitor of barnase) at elevated temperature. Barstar N7C, a mutant constructed in this lab, was linked via a disulfide to a chromatography support. This allows specific elution under mild reducing conditions. Barnase has also been successfully expressed on the surface of phage when attached to the C-terminus of an Fab light chain, with the heavy chain attached to the gene III protein. Again, western blotting analysis was used to verify the success of display. Replacement of the barnase gene with cDNA should allow the successful surface display of cDNA libraries on phage.
Author: Pamela Sears Publisher: ISBN: Category : Languages : en Pages : 5
Book Description
Barnase was successfully displayed on the surface of m13 phage as an N-terminal fusion with coat protein III. The successful display was verified by Western Blotting of purified phage. For selection of a thermostable barnase, the buried residues Ala11, Leu14, Leu20, Ala74, Ile88, Tyr90, and Ile96 were replaced randomly with Phe, Leu, Val, Ile, Ala, Pro, Gly, Ser, or Thr, giving a library of 2x10(exp 6) members. The transformation yield obtained (4x10(exp 6) allows sampling of all possible members. We are currently selecting the phage library on immobilized barstar (the femtomolar inhibitor of barnase) at elevated temperature. Barstar N7C, a mutant constructed in this lab, was linked via a disulfide to a chromatography support. This allows specific elution under mild reducing conditions. Barnase has also been successfully expressed on the surface of phage when attached to the C-terminus of an Fab light chain, with the heavy chain attached to the gene III protein. Again, western blotting analysis was used to verify the success of display. Replacement of the barnase gene with cDNA should allow the successful surface display of cDNA libraries on phage.
Author: Sachdev S. Sidhu Publisher: CRC Press ISBN: 0849359120 Category : Medical Languages : en Pages : 770
Book Description
The first and only guide to showcase the impact of phage display technology on drug discovery, this reference details the theories, principles, and methods impacting the field and demonstrates applications for peptide phage display, protein phage display, and the development of novel antibodies. Highlighting the current and future role of phage dis
Author: Sudhir Paul Publisher: Karger Medical and Scientific Publishers ISBN: 3805570996 Category : Medical Languages : en Pages : 172
Book Description
This volume addresses fundamental questions concerning the immunological genesis of the catalytic activity in antibodies, its relationship with classical antigen binding activity, and the biochemical mechanisms involved in catalysis. The contents reflect three main challenges in the field, i.e. to delineate the biological functions of catalytic antibodies in autoimmune disease; to isolate therapy-grade antibody catalysts with sufficient specificity and turnover to permit rapid removal of microbial and tumor antigens; and to develop immunogens that recruit immature catalyst-producing B cells into the clonal selection pathway and induce adaptive improvements of the catalytic function. Well-edited and up-to-date, this book reviews the current knowledge in the field and explores ways by which natural and engineered catalytic activities can be harnessed for medical applications. It should therefore be of special interest to immunologists, biochemists, biotechnologists, rheumatologists and pathologists.
Author: Frances H. Arnold Publisher: Springer Science & Business Media ISBN: 1592593968 Category : Science Languages : en Pages : 381
Book Description
Directed evolution comprises two distinct steps that are typically applied in an iterative fashion: (1) generating molecular diversity and (2) finding among the ensemble of mutant sequences those proteins that perform the desired fu- tion according to the specified criteria. In many ways, the second step is the most challenging. No matter how cleverly designed or diverse the starting library, without an effective screening strategy the ability to isolate useful clones is severely diminished. The best screens are (1) high throughput, to increase the likelihood that useful clones will be found; (2) sufficiently sen- tive (i. e. , good signal to noise) to allow the isolation of lower activity clones early in evolution; (3) sufficiently reproducible to allow one to find small improvements; (4) robust, which means that the signal afforded by active clones is not dependent on difficult-to-control environmental variables; and, most importantly, (5) sensitive to the desired function. Regarding this last point, almost anyone who has attempted a directed evolution experiment has learned firsthand the truth of the dictum “you get what you screen for. ” The protocols in Directed Enzyme Evolution describe a series of detailed p- cedures of proven utility for directed evolution purposes. The volume begins with several selection strategies for enzyme evolution and continues with assay methods that can be used to screen enzyme libraries. Genetic selections offer the advantage that functional proteins can be isolated from very large libraries s- ply by growing a population of cells under selective conditions.
Author: Benjamin F. Cravatt Publisher: Springer ISBN: 3030111431 Category : Medical Languages : en Pages : 417
Book Description
This volume provides a collection of contemporary perspectives on using activity-based protein profiling (ABPP) for biological discoveries in protein science, microbiology, and immunology. A common theme throughout is the special utility of ABPP to interrogate protein function and small-molecule interactions on a global scale in native biological systems. Each chapter showcases distinct advantages of ABPP applied to diverse protein classes and biological systems. As such, the book offers readers valuable insights into the basic principles of ABPP technology and how to apply this approach to biological questions ranging from the study of post-translational modifications to targeting bacterial effectors in host-pathogen interactions.
Author: Robert Aitken Publisher: Methods in Molecular Biology ISBN: Category : Medical Languages : en Pages : 260
Book Description
In Antibody Phage Display expert researchers explore the latest in this cutting-edge technology, providing an invaluable resource that will guide readers in the design and execution of experiments based around antibody phage display.
Author: John Howl Publisher: CRC Press ISBN: 9780367385736 Category : Languages : en Pages : 503
Book Description
Focuses on Biology, Pharmacology, and Therapeutic Applications The study and diverse applications of bioactive peptides traverse many sub-disciplines within chemistry, biology, physics, and medicine. Answering a long-standing need, Bioactive Peptides focuses on the biology, pharmacology, and therapeutic applications of endogenous peptide mediators and their analogues. Moving peptide science beyond chemical synthesis strategies and into the realms of peptide biology and therapeutics, it presents the overall contribution that peptide science has made to molecular, cellular, and whole organism biology, while also discussing future targets and therapeutic applications. Beneficial for Experts and Novices Alike Part I provides details of bioactive peptides that interact with common drug targets and analyzes some of the most competitive areas of current research worldwide. While it is widely known that mammalian physiological systems utilize bioactive peptides that have yet to be discovered, other animals provide a rich and valuable source of bioactive peptides. This fascinating area of science is the theme of Part II. Parts III and IV investigate the unique bioactivities of various peptides that are ripe for further exploration. This definitive reference also includes: A detailed description and analysis of a broad range of peptides that interact with G protein-coupled receptors, the quantitatively dominant drug target A discussion of non-ribosomal peptides, which hold promise as sources of endogenous mediators Important examples of common methodologies employed to identify, characterize, and further develop bioactive peptides from a range of natural sources With mounting worldwide interest in their therapeutic potential, bioactive peptides--includ
Author: Matthew B. Coppock Publisher: Humana ISBN: 9781071616888 Category : Technology & Engineering Languages : en Pages : 469
Book Description
This volume explores the latest techniques and strategies used to study the field of peptide macrocycles. The chapters in this book ae organized into four parts: macrocycles synthesis, combinational library synthesis and screening, macrocycle characterization, and unique applications. Part One looks at a variety of peptide cyclization methodologies, and Part Two describes methods for the creation of peptide macrocycles libraries and their subsequent screening against biological targets of interest. Part Three discusses the study and characterization of peptide macrocycle-target interactions, and Part Four introduces unique applications for peptide macrocycles, from higher-order structure formation to post-synthetic functional modifications. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and comprehensive, Peptide Macrocycles: Methods and Protocols is a valuable resource for both novice and expert researchers looking to learn more about this developing field.
Author: Sang Yup Lee Publisher: Springer ISBN: 9783319504353 Category : Science Languages : en Pages : 0
Book Description
This book covers the current states of microbial and related technologies that have been developed for the efficient production of chemicals, fuels and materials by integrating strain and enzyme development, fermentation processes, and downstream processes. The book also covers how microbes and microbial products can be employed to facilitate petroleum recovery. Global consequences of bio-based production of chemicals, fuels and materials are also discussed with insights.
Author: Pamela Sears
Author: Pamela Sears
Author: Sachdev S. Sidhu
Author: 
Author: Sudhir Paul
Author: Frances H. Arnold
Author: Benjamin F. Cravatt
Author: Robert Aitken
Author: John Howl
Author: Matthew B. Coppock
Author: Sang Yup Lee