Research Catalog

Title

Proteolytic enzymes : a practical approach / edited by R.J. Beynon, J.S. Bond.

Publication

Oxford ; New York : IRL Press at Oxford University Press, 1989.

Items in the Library & Off-site

Details

Additional Authors

• Beynon, R. J. (Robert J.)
• Bond, Judith S.

Description

xviii, 259 pages : illustrations; 24 cm.

Series Statement

The Practical approach series

Uniform Title

Practical approach series

Subject

• Proteolytic enzymes
• Peptide Hydrolases
• Peptide hydrolases
• Enzymes
• Enzimas
• Peptidases

Bibliography (note)

• Includes bibliographical references.

Contents

• Proteolytic enzymes: nomenclature and classification / Alan J. Barrett -- Terminology and nomenclature -- Peptidase and related terms -- Specificity-subsite terminology -- Catalytic type -- Homology -- The EC classification of peptidases -- What is the EC system? -- What information does the EC list contain? -- When and how can a newly-discovered peptidase be added to the EC list? -- The MEROPS system for the classification of peptidases -- Families -- Clans -- Individual peptidases -- Uses of the MEROPS system -- Steps one might take on discovering a new peptidase -- Purification of proteolytic enzymes / Sherwin Wilk -- Prelude to purification -- Assay -- Initial considerations -- Buffer composition -- Membrane-bound or soluble? -- Membrane-bound enzymes -- Endogenous inhibitors and activators -- General scheme for purification of proteolytic enzymes -- Initial steps -- Intermediate and final steps -- Specialized techniques for proteolytic enzymes -- Peptidyl aldehyde affinity chromatography -- Affinity columns for cysteine proteinases -- Affinity columns for trypsin-like enzymes -- Affinity columns for metalloproteinases -- Other affinity columns -- Optimization of the purification protocol -- Determination of homogeneity -- Purification of the proteasome; EC 3.4.99.46 -- Conclusions: many roads lead to Rome -- Protease assay methods / Gautam Sarath, Michael G. Zeece, Alan R. Penheiter -- Assays with natural substrates -- Endopeptidase assays -- Exopeptidase assays -- Assays with synthetic substrates.
• Endopeptidase and aminopeptidase substrates -- Spectrophotometric assays -- Fluorimetric assays -- Miscellaneous fluorimetric methods -- Carboxypeptidase substrates -- Radiometric assays -- HPLC assays for peptidases -- Capillary electrophoretic analyses of proteases -- Solid-phase protease assays -- Gel electrophoretic methods -- Plate assays -- Miscellaneous solid-phase assays -- Assays for histochemical studies -- Determination of protease mechanism / Ben M. Dunn -- Introduction -- the importance of mechanistic classification -- The serine peptidases -- The cysteine peptidases -- The aspartic peptidases -- The metallopeptidases -- Methods for determining the mechanistic class -- Classification based on 'standard' inhibitors -- Chemical modification/identification -- Site-directed mutagenesis -- Mechanistic distinctions -- intermediates -- Kinetic studies to probe the mechanism in more detail -- Notes on 'ideal' assays -- Kinetic determination of K[subscript m] and k[subscript cat] -- pH dependence of the kinetic parameters -- Solvent deuterium isotope effects -- Transition state analogues and substrate alteration -- Determination of primary specificity of a protease -- Degradation of standard proteins and peptides -- Cleavage of homologous synthetic peptides -- Inhibition of proteolytic enzymes / Guy S. Salvesen, Hideaki Nagase -- Introduction: which inhibitors? -- Principles for using irreversible and reversible inhibitors -- General structure of synthetic inhibitors -- How to live with crossreactivity.
• Practical use of inhibition constants -- Irreversible inhibitors -- Reversible inhibitors -- Non-specific inhibitors -- [alpha]-Macroglobulins -- Peptide aldehydes -- Peptide chloromethyl ketones -- Metal chelators -- Class-specific inhibitors -- Serine proteases -- Cysteine proteases -- Proteasome -- Metalloproteases -- Aspartic proteases -- Inhibitors as active-site titrants -- Cysteine proteases -- Serine proteases -- Protease inhibitors in cell culture -- Suppression of proteolysis -- Therapeutic value of protease inhibitors -- Finding, purification and characterization of natural protease inhibitors / Hideaki Nagase, Guy S. Salvesen -- The meaning of inhibition -- Finding protease inhibitors -- Screening of inhibitors from natural sources -- Finding inhibitors by reverse zymography -- Finding inhibitors from DNA sequences -- Combinatorial protease inhibitors -- Purification of natural protease inhibitors -- Use of the target enzyme as an affinity ligand -- Conventional purification -- Reverse zymography -- Characterization of inhibitors: inhibition kinetics -- The importance of kinetics -- IC[subscript 50] and percentage inhibition -- Practical inhibitor kinetics -- Reversible inhibitors -- Irreversible inhibitors -- Practical applications of protein inhibitors -- Mass spectrometry of proteolysis-derived peptides for protein identification / Bernhard Kuster, Andrej Shevchenko, Matthias Mann -- Mass spectrometry -- Methods of ionization -- Mass analysis -- Tandem mass spectrometry.
• Interrogation of sequence databases using mass spectrometry data -- Choice of protease -- Peptide mass mapping -- Database identification via tandem mass spectrometry -- Analytical strategy for the identification or cloning of proteins using mass spectrometry -- Low-level protein preparation for characterization by mass spectrometry -- Protein visualization -- Proteolytic cleavage of gel separated proteins -- Protein identification by MALDI peptide mass mapping -- Peptide sequencing by nanoelectrospray tandem mass spectrometry -- Towards cloning of proteins using mass spectrometry data -- Post-translationally modified proteins -- Using proteinases for Edman sequence analysis and peptide marking / John Shannon -- The need for digesting proteins to peptides -- Substrate preparation -- Purification techniques -- Sample requirements -- Reduction and alkylation of proteins -- Digestion -- Choice of proteinase -- Conditions for proteolytic digestions -- Digestion of proteins isolated on polyacrylamide gels -- Monitoring a reaction -- Analysis of the proteolytic digestion -- Mass spectrometry -- Electrophoresis -- Ion exchange -- Reverse-phase chromatography -- Data interpretation -- Prevention of unwanted proteolysis / Michael J. North, Robert J. Beynon -- Proteolytic susceptibility of native proteins -- Intrinsic factors determining the susceptibility of proteins to proteolysis -- The influence of other molecules on susceptibility to proteolysis -- Properties of endogenous proteinases -- Identification of proteolysis as a problem.
• Changes in protein properties -- Mimicking an effect with added proteinases -- Checking samples for proteinase activity -- Inhibition of proteinases -- Outline of approaches for reducing proteinase activity -- Suppression of endogenous proteinase activity -- Preventing proteolysis by denaturation -- Use of proteinase inhibitors -- Removal of proteinases -- Choice of starting material -- Cell disruption and fractionation -- Selective removal of proteinases during purification -- Proteolysis of native proteins as a structural probe / Simon Hubbard, Robert J. Beynon -- Factors influencing susceptibility -- Molecular recognition and limited proteolysis -- Prediction of nicksites -- A tool to aid in prediction of sites of limited proteolysis -- Experimental considerations -- Choice of proteinase -- Ratio of proteinase to substrate -- Solution conditions -- Determination of site of proteolysis -- Strategies for limited proteolysis experiments -- Analysis of limited proteolysis data and simulations -- Obtaining quantitative data -- Analysing the data by non-linear curve fitting -- Example reaction schemes -- Simulations and modelling -- Proteases in peptide synthesis / Volker Kasche -- Enzyme properties influencing the product yield and steric purity in protease catalysed peptide synthesis -- Kinetically controlled synthesis -- Equilibrium-controlled peptide synthesis -- Selecting the optimal protease -- Purity of the protease -- P[subscript 1] and P'[subscript 1] specificity.
• Factors controlling the yield and steric purity in the synthesis of a peptide bond with a given enzyme -- Protection of the P'[subscript 1] and activation of the P[subscript 1]-carboxyl group -- pH -- Temperature -- Ionic strength -- Solvent composition -- Peptide synthesis in suspensions with solid product or substrate -- Planning a protease-catalysed synthesis of a peptide bond -- What enzyme? -- Equilibrium-controlled or kinetically controlled synthesis? -- Free or immobilized enzyme? -- Experimental methods for protease-catalysed peptide synthesis -- Enzyme purity and purification -- Enzyme immobilization -- Substrates and buffers -- Monitoring the synthesis; purification of products -- Optimizing the yield -- Proteases in peptide synthesis: limitations and perspectives -- The Schechter and Berger nomenclature for proteinase subsites -- Some commercially available proteases -- Commercially available proteinase inhibitors -- List of suppliers.

ISBN

• 0199630593
• 9780199630592
• 0199630585
• 9780199630585
• 1852211040
• 9781852211042
• 1852211059
• 9781852211059

LCCN

89002113

OCLC

• ocm19458572
• 19458572
• SCSB-14398526

Owning Institutions

Princeton University Library