Enzymes

Enzymes: A Practical Introduction to Structure, Mechanism, and Data Analysis. Robert A. Copeland Copyright  2000 by Wiley-VCH, Inc. ISBNs: 0-471-35929-7 (Hardback); 0-471-22063-9 (Electronic)

ENZYMES
SECOND EDITION

ENZYMES
A Practical Introduction to Structure, Mechanism, and Data Analysis
SECOND EDITION

Robert A. Copeland

A JOHN WILEY & SONS, INC., PUBLICATION
New York /Chichester / Weinheim / Brisbane / Singapore / Toronto

Designations used by companies to distinguish their products are often claimed as trademarks. In all instances where John Wiley & Sons, Inc., is aware of a claim, the product names appear in initial capital or   . Readers, however, should contact the appropriate companies for more complete information regarding trademarks andregistration. Copyright 2000 by Wiley-VCH, Inc. All rights reserved.

No part of this publication may be reproduced, stored in a retrieval system or transmitted in any form or by any means, electronic or mechanical, including uploading, downloading, printing, decompiling, recording or otherwise, except as permitted under Sections 107 or 108 of the 1976 United States Copyright Act, without theprior written permission of the Publisher. Requests to the Publisher for permission should be addressed to the Permissions Department, John Wiley & Sons, Inc., 605 Third Avenue, New York, NY 10158-0012, (212) 850-6011, fax (212) 850-6008, E-Mail: PERMREQWILEY.COM. This publication is designed to provide accurate and authoritative information in regard to the subject matter covered. It is sold withthe understanding that the publisher is not engaged in rendering professional services. If professional advice or other expert assistance is required, the services of a competent professional person should be sought. ISBN 0-471-22063-9 This title is also available in print as ISBN 0-471-35929-7. For more information about Wiley products, visit our web site at www.Wiley.com.

To Clyde Worthen forteaching me all the important lessons: arigato sensei. And to Theodore (Doc) Janner for stoking the fire.

CONTENTS
Preface Acknowledgments Preface to the First Edition 1 A Brief History of Enzymology 1.1 Enzymes in Antiquity / 2 1.2 Early Enzymology / 3 1.3 The Development of Mechanistic Enzymology / 4 1.4 Studies of Enzyme Structure / 5 1.5 Enzymology Today / 7 1.6 Summary / 8 References andFurther Reading / 10 2 Chemical Bonds and Reactions in Biochemistry 2.1 Atomic and Molecular Orbitals / 11 2.2 Thermodynamics of Chemical Reactions / 23 2.3 Acid—Base Chemistry / 29 2.4 Noncovalent Interactions in Reversible Binding / 32 2.5 Rates of Chemical Reactions / 35 2.6 Summary / 41 References and Further Reading / 41 3 Structural Components of Enzymes 3.1 3.2 3.3 3.4 3.5 The Amino Acids/ 42 The Peptide Bond / 53 Amino Acid Sequence or Primary Structure / 55 Secondary Structure / 57 Tertiary Structure / 62
vii

xi xiii xv 1

11

42

viii

CONTENTS

3.6 Subunits and Quaternary Structure / 65 3.7 Cofactors in Enzymes / 68 3.8 Summary / 71 References and Further Reading / 74 4 Protein‒Ligand Binding Equilibria The Equilibrium Dissociation Constant, K / 76 The KineticApproach to Equilibrium / 78 Binding Measurements at Equilibrium / 80 Graphic Analysis of Equilibrium Ligand Binding Data / 88 Equilibrium Binding with Ligand Depletion (Tight Binding Interactions) / 94 4.6 Competition Among Ligands for a Common Binding Site / 95 4.7 Experimental Methods for Measuring Ligand Binding / 96 4.8 Summary / 107 References and Further Reading / 108 4.1 4.2 4.3 4.4 4.5 5Kinetics of Single-Substrate Enzyme Reactions 5.1 The Time Course of Enzymatic Reactions / 109 5.2 Effects of Substrate Concentration on Velocity / 111 5.3 The Rapid Equilibrium Model of Enzyme Kinetics / 113 5.4 The Steady State Model of Enzyme Kinetics / 115 5.5 The Significance of k and K / 120 
5.6 Experimental Measurement of k and K / 124 
5.7 Other Linear Transformations of Enzyme...