Glutathione. Proceedings of the Symposium Held at Ridgefield, Connecticut, November, 1953

342 Pages·1954·22.408 MB·English

GLUTATHIONE
Proceedings of the Symposium
Held at Ridgefield, Connecticut
November, 1953
EDITORS
S. Colowick D. R. Schwarz
A. Lazarow E. Stadtman
E. Racker H. Waeisch
J 9 54
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N EW Y O RK
Copyright 1954 by
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Library of Congress Catalog Card Number: 54-11062
PRINTED IN THE UNITED STATES OF AMERICA
List of Contributors
MARK D. ALTSCHULE, McLean Hospital, Waverly, Massachusetts
C. B. ANFINSEN, JR., National Institutes of Health, Bethesda, Maryland
L. ASTRACHAN, Johns Hopkins University, Baltimore, Maryland
RUSSELL J. BARRNETT, Harvard Medical School, Boston, Massachusetts
E. S. G. BARRON, The University of Chicago, Chicago, Illinois
REINHOLD BENESCH, State University of Iowa, Iowa City, Iowa
RUTH E. BENESCH, State University of Iowa, Iowa City, Iowa
F. BINKLEY, Emory University, Georgia
KONRAD BLOCH, The University of Chicago, Chicago, Illinois*
LOUISE B. BRADLEY, National Institutes of Health, Bethesda, Maryland
GEORGE BRECHER, Naval Medical Research Institute, Bethesda, Maryland
MELVIN CALVIN, University of California, Berkeley, California
WILLIAM H. CHAPMAN, Naval Medical Research Institute, Bethesda, Maryland
H. T. CLARKE, Columbia University, New York, Ν. Y.
SIDNEY P. COLOWICK, Johns Hopkins University, Baltimore, Maryland
ROBERT A. CONRAD, Naval Medical Research Institute, Bethesda, Maryland
ERIC E. CONN, University of California, Berkeley, California
G. E. CONNELL, University of Toronto, Toronto, Canada
E. P. CRONKITE, Naval Medical Research Institute, Bethesda, Maryland
ZACHARIAS DISCHE, Columbia University, New York, Ν. Y.
G. H. DIXON, University of Toronto, Toronto, Canada
J. W. DUBNOFF, California Institute of Technology, Pasadena, California
J. W. DUCKWORTH, Naval Medical Research Institute, Bethesda, Maryland
SALLY W. EDWARDS, New England Deaconess Hospital, Boston, Massachusetts
P. J. FODOR, Flower and Fifth Avenue Hospital, New York, Ν. Y.
J. S. FRUTON, Yale University, New Haven, Connecticut
REBECCA GERSCHMAN, University of Rochester, Rochester, New York
ROSE MARIE GONCZ, McLean Hospital, Waverly, Massachusetts
G. R. GREENBERG, Western Reserve University, Cleveland, Ohio
MARCIA GUTCHO, Schwarz Laboratories, Inc. 230 Washington Street, Mount Vernon,
New York
C. S. HANES, University of Toronto, Toronto, Canada
L. HELLERMAN, Johns Hopkins University, Baltimore, Maryland
DOROTHY H. HENNEMAN, McLean Hospital, Waverly, Massachusetts
F. J. R. HIRD, University of Melbourne, Victoria, Australia
SIDNEY INGBAR, Thorndike Memorial Laboratory, Boston, Massachusetts
W. WAYNE KIELLEY, National Institutes of Health, Bethesda, Maryland
W. E. KNOX, New England Deaconess Hospital, Boston, Massachusetts
M. E. KRAHL, University of Chicago, Chicago, Illinois
Louis LAUFER, Schwarz Laboratories, Inc. 230 Washington Street, Mount Vernon,
New York
ARNOLD LAZAROW, Western Reserve University, Cleveland, Ohiof
* Present address: Department of Chemistry, Harvard University, Cambridge, Massachusetts
t Present address: Department of Anatomy, University of Minnesota, Minneapolis, Minnesota
vi LIST OF CONTRIBUTORS
L. LEVIN, National Science Foundation, Washington, D. C.
I. M. LONDON, Columbia University, New York, Ν. Y.
DANIEL MAZIA, University of California, Berkeley, California
HARVEY M. PATT, Argonne National Laboratory, Lemont, Illinois
J. W. PATTERSON, Department of Anatomy, Western Reserve University, Cleveland,
Ohio
E. RACKER, Yale University, New Haven, Connecticut*
WILLIAM I. ROGERS, State University of Iowa, Iowa City, Iowa
ARNOLD M. SELIGMAN, Harvard Medical School, Boston, Massachusettsf
JOHN E. SNOKE, University of Chicago, Chicago, Illinois
E. R. STADTMAN, National Institutes of Health, Bethesda, Maryland
JAKOB A. STEKOL, Lankenau Hospital, Philadelphia, Pennsylvania
H. J. STRECKER, New York Psychiatric Institute, New York, Ν. Y.
BIRGIT VENNESLAND, University of Chicago, Chicago, Illinois
E. P. VOLLMER, Naval Medical Research Institute, Bethesda, Maryland
HEINRICH WAELSCH, New York Psychiatric Institute, New York, Ν. Y.
G. WALD, Harvard University, Cambridge, Massachusetts
THEODOR WIELAND, Institut f. organische Chemie, Frankfurt/Main, Germany
* Present address: Public Health Research Institute, New York, Ν. Y.
t Present address: Department of Surgery, Sinai Hospital, Baltimore, Maryland
Preface
It is true ... to say that in scientific borderlands not only are facts
gathered that are often new in kind, but it is in these regions that wholly
new concepts arise.
F. G. HOPKINS
Linacre Lecture, 1988
There are many avenues whereby communication between scientists
occurs. It is almost axiomatic that one of the most desirable means for
promotion of communication is direct discussion between the interested
persons. Intimate contact between investigators working in the same field
provides a stimulus to all concerned. To this end, small conferences and
symposia, attended by the best informed persons working in a particular
scientific field, are an extremely valuable aid to research. Not only do such
conferences permit dissemination and consideration of newly obtained
information but they also facilitate exchange of ideas, hypotheses, and
theories as well as critical examination of the state of knowledge in the
field under consideration. Such evaluation may be expected to aid research
progress by emphasizing those scientific areas where more knowledge is
urgently needed as well as by pointing out others in which further research
does not appear profitable at the present time. A very valuable by-product
of such conferences and symposia is the publication of a complete tran-
script of the proceedings which permits those who have not attended the
meeting, including those who have not yet entered the particular scientific
field, to benefit from the conference and from the ideas exchanged there.
Although the need for a conference in a special field may manifest itself
in a variety of ways, its desirability is most reassuringly demonstrated by
the spontaneous development of interest among the investigators actually
engaged in research in the particular scientific field. The present Sym-
posium on Glutathione is an excellent example of a conference originating
in this way.
The National Science Foundation and the Office of Naval Research are
pleased to assist in the organization and support of such conferences. We
welcome the opportunity to help advance scientific progress in this way as
in many others.
ALAN T. WATERMAN, Director, National Science Foundation
C. M. BOLSTER, Rear Admiral, USN, Chief of Naval Research
Acknowledgment
We wish to express our gratitude to the National Science Foundation
and the Office of Naval Research for the joint support which made this
Symposium possible. Dr. Louis Levin of the National Science Foundation,
and Mr. Leo A. Shinn, Dr. Freeman Quimby, Dr. Eli Goldsmith and Mrs.
Elizabeth Kelly, of the Office of Naval Research, deserve individual men-
tion for many helpful suggestions.
The College of Physicians and Surgeons of Columbia University served
as the administrating agency. We wish to thank Dean Willard C. Rappleye
for accepting this responsibility, and are particularly indebted to Mr.
Bernis D. Moss, Jr., Business Officer of the College, for his efficient han-
dling of administrative details.
Mrs. Rose Rubino and Miss Josephine Shyers, who transcribed the dis-
cussions during the meetings, merit special acknowledgment for a difficult
task well done.
THE EDITORS
Mercaptans and Disulfides : Some Physics, Chemistry, and
Speculation1
MELVIN CALVIN
I. Introduction 3
II. Physics 6
III. General Chemistry—Acidity 8
IV. General Chemistry—Addition Reactions 11
V. General Chemistry—Mercaptides and Displacement Reactions 14
VI. General Chemistry—Oxidation 15
VII. General Chemistry—Disulfides 15
VIII. Oxidation Potential 19
IX. Application to the Structure of Glutathione 21
I. Introduction
The subject of this conference is given as "Glutathione." This is a
very comprehensive subject. However, it does have some limitations, and
the primary part of that subject to which this paper is addressed is the
chemistry of the SH and SS systems such as might conceivably be involved
in glutathione chemistry.
The generally accepted formula for glutathione is 7-glutamylcysteinyl-
glycine, which is really a diamide, having the structure
Ο Ο
II II
~0 C—CH—CH2CH2—C—NH—CH—C—NU—CH —CO2H
2 2
I I
+NH CH
3 2
I
SH
7-glutamyl cysteinyl glycine
One of the major points of interest in the chemistry of glutathione has been
the chemistry of the mercaptan group. More recently, there has been an
additional interest developed in the possibility of glutathione being a com-
mon intermediate in the synthesis of all peptide links, particularly in view
of the existence of enzyme systems which transfer this 7-glutamyl residue
to a whole variety of other amino acids. However, this is not the part of
the subject to which I will be addressed. We will limit ourselves, then, to
a discussion of sulfur chemistry.
1 The work described in this paper was sponsored by the U.S. Atomic Energy
Commission.
3
4 PROPERTIES AND ORGANIC CHEMISTRY

1 "
[phone]-S0-OH 2 Ο II 0-S-O-R II Ο Potentials,
SSS . - - n
ANALOGUES listed.) +5 HHH222 -0.17 -0.17 -0.17 7) 1 | .57 -0M HS0e 22 R R Ο Ο II II RO-S-S-OR II II Ο Ο er in "Oxidatio
TABLE I« SOME OXIDATION STATES OF SULFUR COMPOUNDS AND THEIR ORGANIC (Only compounds with one or two sulfur atoms in the molecule are 0 -1 +4 +2 +3 +1 (-0.5) (-0.4) s[hso] HS0HS0HS0HS0223 23 23 22 1 r -0.14 (-0.4) 1 1 1 0.25 (-0.003) 1 1 1 (-0.-0.40 -0.40 -0.40 -0.40 1 1 11 -0.51 1 -01 _ I -HS0 HS0HS03 SHS04 22222 222222 -0.88| -0.08 > -0.45 Ο Ο II II R-S-OH lR-0-S-OR R-S-OH Ο Ο II II R-S-O-R R-S-O-R R-S-O-R 00 %Ο -0.23 -0.9 II R-S-R R-S-R II R-S-0-H II Ο II ιΟ Ο Ο Ο Ο Ο / \ II 11 R-S-S-OHlR-0-S-S-O-R -S-S-RlR-S S-RlR-S-SH II Ο Ο O Ο II R-S-S-R II R-S-S-R II Ο +tials are for liVH, and the sign is according to the convention of W. LatimHall, New York, 1938.
H R n
-2 HS 2 1 - -0.11 < R-S-H R-S-R ° PotePrentice-
MERCAPTANS AND DISULFIDES 5
In order to do this properly, one should have a more general view of the
nature of sulfur chemistry, both inorganic and organic. For this purpose,
I have prepared a table (Table I) showing the various oxidation levels of
sulfur, ranging from —2 to +6. Both inorganic sulfur compounds and
some of their organic analogues are listed. They are arranged in two rows;
the molecules with one atom of sulfur in them and the molecules containing
two atoms of sulfur. There exist, of course, others with higher numbers,
but for the moment we will not be concerned with them. Included also
are a number of oxidation potentials in which we might be interested, con-
necting various inorganic species. It may be possible to use those inor-
ganic redox values to estimate what the corresponding organic compound
would have, since only three of these latter are listed and of the three only
the mercaptan-disulfide («—2 «-> —1) system has had any direct measure-
ments attempted upon it. These will be discussed a bit more later. It is
perhaps worth pointing out that for those compounds containing more than
one sulfur atom, the oxidation number listed is the average one taken over
all the sulfur atoms in the molecule. If they are separated in a nonredox
process (such as hydrolysis) there may be a marked dismutation of oxida-
tion number and reduction potential. This is, of course, especially true of
molecules in an odd average oxidation number, i.e.,
R—S—S—R + H 0 -> R—SH + R—S—OH
2
-1 -2 0
R—S—S—R + H 0 -» R—S—OH + RSH
2
II II
ο ο
0 +2 -2
Such a process of average oxidation, usually with lower potential require-
ment followed by an internal rearrangement of redox potential, may pro-
vide a route in biological systems for electron transfer through otherwise
prohibitive (direct) potential barriers.
Now the particular interest we have in the present discussion is in the
organic compounds containing sulfur corresponding to the two levels —2
and — 1 ; that is, a mercaptan or a thioether, and a dialkyl disulfide, or to
complete the analogy an alkyl hydrogen persulfide.
R—SH R—S—S—Η
or or
R—S—R R—S—S—R
-2 -1
Of these, by far the most important for glutathione chemistry, so far, is
the alkyl mercaptan-dialkyl disulfide system.
6 PROPERTIES AND ORGANIC CHEMISTRY
II. Physics
The first thing I thought we would discuss is the physical evidence about
the structure of these two groups. What do we know about such things
as the thermodynamics of mercaptans and the disulfides? What do we
know of the geometrical arrangement of the bonds around these groups?
The distances are known, some from spectroscopic data and some from
crystal or gas diffraction data. They are given in Table II. The bond
energy can be computed. Before we go into the bond energy, let us say a
little about the angles and geometry of the sulfides. The bond angle
around a divalent sulfur is somewhat over 90 deg. as far as it has been
determined. The bonds in divalent sulfur might be considered as essen-
tially p-bonds, the d-orbital playing relatively little part, and in this sense
it is very similar to the bonding in oxygen compounds. For that reason,
then, two bonds for the divalent sulfur atoms are roughly at 90 deg. (it
varies with the particular substituent, being in general somewhat greater).
The geometry of the disulfide-containing molecule is very interesting, the
reason being that in this case one has two such sulfur atoms, presumably
in both cases bonded by ordinary p-electron pairs (to a first approxima-
tion). The s pair is spherically distributed about the nucleus, and the ρ
pairs are on the 90-deg. axes. The result is that in a disulfide the distribu-
tion leads to a very interesting geometry. An attempt to show this is made
in Fig. 1. The spatial configurations (dimethylsulfide), for example, are
about as shown. First the S—S—C bond angle (7) is about 107 deg., but
the important and interesting thing is that the dihedral angle (δ) between
the two S—C bonds is very nearly 90 deg. There is a restriction to the
rotation about the S—S link amounting (at its maximum) to at least 10
Kcal., and probably nearer 20 Kcal, (from heat capacity measurements),
which is a very high restriction for what appears to be a single bond between
TABLE II
a
SOME BOND ENERGIES (E) AND DISTANCES (d)
Ε (Kcal.) d(A.)
O--H 110 0.957
s- -H 82 1.345
c--O 74 1.44
c--s 52 1.81
H--H 104 0.749
s- -S 50 2.04
c--c 65 1.55
c= =0 150
a For more complete lists, see M. L. Huggins, Am. Chem. Soc. 75, 4123 (1952);
K. S. Pitzer, "Quantum Chemistry," Prentice-Hall, New York, 1953.