Solvent-Dependent Flexibility of Proteins and Principles of Their Function

301 Pages·1985·10.445 MB·English

by  Alex I. Käiväräinen (auth.), Alex I. Käiväräinen (eds.)

Solvent-Dependent Flexibility of Proteins
and Principles of Their Function
Advances in Inclusion Science
Alex I. Kaivarainen
Institute of Biology, U.S.S.R. Academy of Sciences
Solvent-Dependent Flexibility
of Proteins
and Principles of Their Function
D. Reidel Publishing Company
lI-..
A MEMBER OF THE KLUWER ACADEMIC PUBLISHERS GROUP "
Dordrecht / Boston / Lancaster
Library of Congress Cataloging in Publication Data
Kiiiviiriiinen, A. I. (Aleksandr Ivanovich)
Solvent-dependent flexibility of proteins and principles of their function.
(Advances in inclusion science)
Translation of: Dinamicheskoe povedenie belkov v vodnoi srede i ikh funktsii.
Bibliography: pp.
Includes index.
1. Proteins. 2. Water of hydration. I. Title. II. Series.
QP551.K4913 1983 574.19'245 83-19121
ISBN-13: 978-94-010-8798-8 e-ISBN-13: 978-94-009-5197-6
DOl: 10.1007/978-94-009-5197-6
Published by D. Reidel Publishing Company,
P.O. Box 17, 3300 AA Dordrecht, Holland
Sold and distributed in the U.S.A. and Canada
by Kluwer Academic Publishers,
190 Old Derby Street, Hingham, MA 02043, U.S.A.
In all other countries, sold and distributed
by Kluwer Academic Publishers Group,
P.O. Box 322, 3300 AH Dordrecht, Holland.
All Rights Reserved
© 1985 by D. Reidel Publishing Company, Dordrecht, Holland
Softcover reprint ofthe hardcover 1s t edition 1985
No part of the material protected by this copyright notice may be reproduced or utilized in any form
or by any means, electronic or mechanical including photocopying, recording or by any information
storage and retrieval system, without written permission from the copyright owner
Table of Contents
Foreword ix
Introduction xi
Chapter 1 / The Properties of Water
1.1. Pure Water
1.2. Water in Model Systems 8
Chapter 2 / Principles and Implications of the Dynamic Model of Behavior of
Proteins in Aqueous Media 14
2.1. Description of the Properties of Proteins in Terms of the Dynamic Model 14
2.2. A Description of the Properties of Membranes in Terms of the Dynamic
Model 22
Chapter 3 / Conformational Mobility of Proteins 27
3.1. Studying Hydrated Biopolymers by IR Spectroscopy 29
3.2. Fundamental Principles of Hydrogen Exchange Method 30
3.3. Results Obtained by Hydrogen Exchange and Proteolytic Studies 31
3.4. Fundamental Principles and Applications of the NMR Method 33
3.5. Gamma-Resonance Spectroscopy and its Applications 37
3.6. The Method of Spin Label and its Application 40
3.7. Application of Luminescence to the Study of Protein Dynamics 48
3.8. Results of X-Ray Diffraction Studies of Protein Mobility 53
Chapter 4/ Interaction Between Proteins and Water 57
4.1. Hydration of Biopolymers and Methods of its Study 57
4.2. Protein Cavities (Clefts) and their Interaction with Water 64
4.3. The Dynamics of Water-Protein Interaction 70
Chapter 5 / Effect of Perturbing Agents on the Dynamic Properties of Proteins 80
5.1. Changes in the Conformation and Stability of Proteins Brought about by
Perturbing Agents 84
vi Table of Contents
5.2. Effect of Temperature on Protein Dynamics and on the Interaction
between the Protein and the Surrounding Solvent 97
5.3. Ultrasonic Action 109
Chapter 6 / Dynamic Model of Association and Dissociation of Specific Complexes 111
6.1. Effect of Complex Formation on the Stability of the Protein 118
6.2. Effect of Complex Formation on Dynamic Properties of Proteins 119
6.3. Immunoglobulins 120
6.4. Cytochrome c 163
6.5. Myoglobulin 168
6.6. Hemoglobin 174
Chapter 7 / Dynamic Model of Protein Behavior and Mechanism of Enzymatic
Function 198
7.1. A Possible Connection between the Kinetic and Conformational
Properties of Enzymes 198
7.2. Comparison between the Dynamic Model of Enzymatic Activity and
Other Models of Enzymatic Catalysis 205
7.3. Relaxational Properties of Enzymes 211
7.4. Effect of Perturbing Agents on Enzymatic Activity 214
7.5. Action of D2 0 on Enzymes 217
7.6. Glyceraldehyde-3-Phosphate Dehydrogenase 217
7.7. T ransamina se 221
7.S. Myosin 223
7.9. Lysozyme 226
7.10. a-Chymotrypsin and Pepsin 229
7.11. Study of Reactions Involving Papain at Low Temperature 233
7.12. Allosteric Enzymes 234
Chapter 8 / Interactions between Macromolecules of Different Types and Proteins
and Cells in Aqueous Media 238
8.1. Effect of Polymers on the Flexibility of Antibodies in Complex with
Spin-labeled Hapten at Different Temperatures 239
8.2. Effect of Proteins on the Flexibility of Antibodies and Oxyhemoglobin 240
8.3. Effect of Thermality Induced Transitions of Human Serum Albumin on
the Mobility of the Subunits of Spin-labeled Oxyhemoglobin 242
8.4. Effect of Polymers and Proteins in Different Ligand States of Active Sites
on the Properties of the Solvent 243
8.5. Interaction between Serum Proteins and Cells 247
Table of Contents vii
Conclusion 254
References 258
Index of Authors 281
Index of Subjects 286
Foreword
Molecular biology has now advanced to the point where it is no longer possible to give a
complete review of the available data on the conformational features of proteins. New
data keep streaming in, and there is obviously an urgent need for some sort of general
treatment of the subject. A systematic treatment of the large amount of data obtained by
a great variety of methods on a great variety of objects must be based on the use of
models; these should be as simple as possible, should conform to well-established scien
tific laws, and at the same time be sufficiently flexible. The validity of the models
finally arrived at is then confirmed or otherwise by testing the conclusions arrived at with
their aid by means of experiment.
After a suitable model has been adopted, it can be used in analyzing the experimental
data. Such an analysis may result in one of three possible situations: neither the experi
mental results nor their interpretation contradict the proposed model; the experimental
results do not contradict the proposed model but their interpretation by the authors
does; finally, both the experimental results and their interpretation may be found to be
incompatible with the fundamental principles underlying the proposed model.
The first situation is clearly the most desirable, and presents no difficulties. If the sec
ond situation obtains, we would do well to recall the wise words of Leonardo da Vinci:
"Experiments are never wwng; it is only opinions which may be ... " The third situation
is the most serious: if the experimental setup does not give rise to any systematic errors,
the model must be modified or even discarded altogether. Here, one firm 'against' out
weights any number of 'fors'.
This book will consider three models, each succeeding model more complex than the
previous one: the dynamic equilibrium model of behavior of proteins in water; the dyna
mic association-dissociation model of specific complex compounds; and the dynamic
model of enzymatic activity.
The principles of these models derive from relatively simple systems, which are taken
as examples. The problems involved in the interaction of proteins with water form one of
the principal subjects of this book. A number of realistic examples are employed to de
monstrate the validity of the model, and in order to show that different experimental
data can be reduced to a common denominator by its use. As a result, an altogether novel
interpretation of familiar facts becomes possible.
It is my pleasant duty to thank L.A. Blumenfeld, G.1. Likhtenstein, Yu. I. Khurgin,
V.I. Lobyshev, S.I. Aksenov and A.V. Filatov for their assistance and for the comments
they offered on the manuscript.
ix
Introduction
The properties of proteins, in the form of results obtained by biochemical and physico
chemical studies, form the subject of a very large number of publications. Interest in the
connection between the structure and the biological functions of proteins is steadily
growing. The concept of the active protein as a cooperative system with definite degrees
of freedom has now become firmly established, and most experts now believe that there
are certain general principles which govern protein activity.
The major advances of molecular biology are now apparent to all; nevertheless, the
mechanism of functional activity is not yet fully understood, not even for a single pro
tein. Attempts to solve this problem, including the application of the powerful tool of
X-ray diffraction analysis, have so far failed, even though detailed information about
the initial (ligand-free) and final (ligand-bonded) states of the macromolecule was avail
able. Even if it is assumed that the conformational states of the molecule in solution and
in the crystal" are identical - an assumption which is itself open to doubt - we are still un
able to effect an experimental follow-up of the complicated pathways of the structural
rearrangements involved in the conversion from the initial to the final state or to grasp
the meaning of their particular sequence. Likely methods of approach to this problem in
futUre will most probably be dynamic such as NMR, EPR, fluorescence methods, deute
rium exchange techniques, stopped flow method, temperature jump method, etc.
During the past few years the principal scientific interest has shifted to the dynamic
aspects of protein physics. These questions are intimately connected with the problems
involved in the hydration of biopolymers, which are now being intensively studied.
A detailed treatment of the nature of the interaction between proteins and water,
including correlation times and percentage contents of the different fractions is given in
numerous studies, with the main stress on the role of water in the mobility of the protein
matrix. However, practically nothing has been published on the conformational changes
taking place in the solvate hull as a result of conformational rearrangements of the pro
tein, except possibly about its changed extent of hydration. Only very few workers con
sider the sorbed water to be an active participant in the structural changes taking place in
the macromolecule.
The principal subject of this book is the treatment of the dynamic properties of pro
teins as related to the properties of their solvate hulls, and of the general relationships
governing the interaction of proteins with water. It will be shown that the available data
can be readily fitted into our dynamic model of behavior of proteins in water, without
resulting in any inconsistencies. The model offers a systematic explanation for the results
of studies conducted on a given object by several different methods, and makes it possible
to consider a variety of different phenomena from a uniform point of view. Several non
trivial implications of the model were confirmed by series of specially conducted ex
periments. Cortxersely, no facts could be discovered which would contradict any funda-
xi
xii Introduction
mental conception, even though every effort was made at an objective approach to this
problem.
Our model introduces one novel concept - the idea of 'clusterophilic' interaction be
tween protein cavities and water. The concept follows from the hypothesis that protein
cavities with their active sites assume, in one of their states, a special structure (geometry,
amino acid composition, etc.), in which the water molecules which have entered the
cavity arrange themselves into an ordered, cooperative 'cluster', since this is thermodyna
mically favored. The lifetime of such 'clusters' is not more than about 10-8 second; it
varies with the properties of the protein and with external conditions, and is several or
ders longer than that of hydrogen bonds in homogeneous liquid water (10-11 - 10-12
sec). If, for any reason, the water has lost its capacity to form clusters, its interaction
with the cavity is thermodynamically less favored; as a result, the stability of the protein
cavity decreases, and the cavity tends to assume another structure which is stable by vir
tue of its hydrophobic properties. The former state will be referred to as the 'open' B
state, while the latter will be denoted as the 'closed' A-state. It is probable that this
adaptability of the properties of proteins to those of water is the result of a prolonged
biological evolution, in the course of which the macromolecules have 'learned' to utilize
the phase transitions of water to regulate their own confonnation and stability (Kaivar
iiinen, 1975b).
The idea that water may have affected the evolution of protein properties was also
pu t forward at an earlier date, though in a less specific form : "Proteins are a machine,
which has been programmed by evolution to undergo chemical reactions with liquid
water" (Lamri and Biltonen, 1973). According to these workers, this is realized by way of
fine changes, which involve both the geometry of the proteins and the state of the sol
vent, including the adaptation of the conformations of the protein and the water to each
other. They identify this process as the swelling of proteins, which is accompanied by free
exchange of water. At 25°C the enthalpils required to produce a hole of the approximate
size of one H 0 molecule in water, in short-chain aliphatic alcohols and in pure aliphatic
2
hydrocarbons are about 84,29 and 8.5 kilo-joules respectively*. It is thought, according
ly, that cavities in proteins will appear mostly at the concentration sites of hydrophobic
residues, and at the sites of contact between the hydrophobic and the polar groups.
The problem of the effect of water on biological systems was actively studied by
Drost-Hansen (1970), who explained the observed anomalies in the variations of various
biological processes in terms of polymorphic transitions between the different structures
of water. A study of a large number of data on temperature anomalies led this worker to
specify four typical transition temperatures: 15,30,45 and 60°C. However, the attempt
by O'Neil and Adami (1969) at confirming temperature-dependent isomorphism of water
was unsuccessful.
Following the work of Frank and Evans (1945), who pointed out that nonpolar com
pounds tend to stabilize the adjacent water molecules, many hypotheses on the role of
'ice-like' water in biological systems have been advanced. Klotz (1964) suggested that
water reacts with nonpolar side chains of the protein to form crystalline hydrates and
* All physical units in this book are SI units.