Table Of ContentBayer-Symposium V
Proteinase Inhibitors
Proceedings of the 2nd International
Research Conference
Edited by
H. Fritz, H. Tschesche, L. J. Greene,
and E. Truscheit
Springer-Verlag New York· Heidelberg· Berlin 1974
Dr. HANS FRITZ, Institut flir Klinische Chemie und Klinische Biochemie der Universitat
Miinchen, 8000 Munich 2, Fed. Rep. Germany
Dr. HARALD TSCHESCHE, Organisch-Chemisches Laboratorium der Technischen
Universitat Miinchen, 8000 Munich 2, Fed. Rep. Germany
Dr. LEWIS 1. GREENE, Brookhaven National Laboratory, Department of Biology,
Upton, NY 11973, USA
Dr. ERNST TRUSCHEIT (Editor-in-Chief), Biochemisches Laboratorium der Bayer AG,
5600 Wuppertall, Fed. Rep. Germany
The editors are indebted to Miss M. L. CHAMBERLAIN and Dr. W. WINGENDER
for their valuable assistance and in particular to Mrs. B. NAGEL
for careful proof-reading and assistance in the preparation of the subject index
Bayer-Symposium V
(2nd International Research Conference on Proteinase Inhibitors)
held at Grosse Ledder near Cologne, Fed. Rep. Germany
October 16th-20th, 1973
With 410 Figures. A stereo viewer for the
stereo pair figures on pages
476,488,489,493,502,504,
505,507,509,515, and 519
is attached inside the back cover
ISBN-13: 978-3-642-87968-5 e-ISBN-13: 978-3-642-87966-1
DOl: 10.1007/978-3-642-87966-1
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© by Springer-Verlag Berlin· Heidelberg 1974. Library of Congress Catalog Card Number 74-6880.
Softcover reprint of the hardcover 1st edition 1974
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Preface
These proceedings contain the papers presented at the Bayer-Symposium V
which was held in Grosse Ledder on October 16-21, 1973 and which was
simultaneously the Second International Research Conference on Proteinase
Inhibitors.
The cordial atmosphere, the frank discussions and the profitable exchange of
ideas during the First International Research Conference on Proteinase Inhibitors
which was held on November 4--6, 1970 in Munich were a stimulant for us to
consider holding a Second Conference especially as the steadily increasing research
activities in this and related fields called for a new assessment of recent efforts in
proteinase inhibitor research.
In the three years since the First Conference was held, this field has developed
particularly in the areas of molecular aspects of protein-protein interactions and in
the extension to inhibitors of nonserine proteinases and peptidases. In planning
this meeting the Organizing Committee was faced with the problem of selecting a
limited number of participants and we decided that it was appropriate and
necessary to focus in detail on the chemistry of proteinase inhibitors, their interac
tions with a variety of enzymes and also to invite more general papers to survey
the physiological and clinical aspects of the subject which we felt would be of
importance in the future.
The organizers have been aware of the fact that not all of the more recent
aspects could be considered. The number of participants and papers presented
has more than doubled since the First Conference, despite the careful selection of
the contributions, thus the size of the Conference Proceedings has considerably
increased from about 300 to 800 pages. This explains why it was unfortunately
impossible in this volume to include the extended, lively and extremely stimulating
discussions which were enjoyed by the participants during the Conference.
The contributions of Dr. G. FEINSTEIN, University of Tel-Aviv, who was
unfortunately prevented from attending have, with the permission of all partici
pants, been included in this volume without having been presented at the
Conference. We are particularly grateful to Professor M. LASKOWSKI,Jr. for his
excellent resume of the symposium.
The editors are greatly indebted to all contributors and to all those who gave
assistance without which the production of this volume would not have been
accomplished. The preface permits the editors to acknowledge the conscientious
work of the publisher.
It is with great respect and affection that the organizers of the symposium, on
behalf of all participants and the Bayer AG, dedicate this volume to Professor
MICHAEL LASKOWSKI, Sr. for his significant, extensive research in the area of
proteinase inhibitors and for his example to us both as a man and as a scientist.
HANS FRITZ . HARALD TSCHESCHE . LEWIS J. GREENE . ERNST TRUSCHEIT
*
Contents
A Turbulent Life: M. LASKOWSKI, Sr. 3
Welcome: E. TRUSCHEIT 11
Section I
Plasma Proteinase Inhibitors
Biochemistry of Proteinase Inhibitors from Human Plasma: A Review of
Recent Development: N. HEIMBURGER. With 2 Figures .. . . . .. 14
Purification, Properties, and Composition of oc r Trypsin Inhibitor from
Human Plasma: L. F. KRESS and M. Laskowski, Sr. With 7 Figures 23
Properties of Human ocl-Antitrypsin:J. Travis, D. JOHNSON, and R. PANNELL.
With 8 Figures ........................ 31
Human oci-Antitrypsin Phenotypes: F. KOPPERS. With 3 Figures 40
Isolation and Fragmentation of ocrAntitrypsin: J.-O. Jeppsson and c.-B.
LAURELL. With 3 Figures .................... 47
The Interaction of Human Cationic Trypsin and Chymotrypsin II with
Human Serum Inhibitors: J. Bieth, M. AUBRY, and J. TRAVIS. With
7 Figures . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Studies on the Quaternary Structure of Human Serum OCr Macroglobulin :
R. C. Roberts, W. A. RIESEN, and P. K. HALL. With 5 Figures . . . . . 63
The Unique Nature of the Interaction of oc2-Macroglobulin with Proteinases:
A.J.Barrett, P. M. STARKEY, and E. A. MUNN. With 2 Figures . . . . 72
Biology and Pathology of Plasma Proteinase Inhibitors: M.Steinbuch and
R. AUDRAN. With 4 Figures . . . . . . . . . . . . . . . . . . . 78
Interaction between Endogenous Proteases and Plasma Protease Inhibitors
in vitro and in vivo: K. OHLSSON. With 11 Figures . . . . . . . . . 96
Plasma Clearance of Human Antiproteinase/Proteinase Complexes: C. BLA-
TRIX, J. ISRAEL, R. AUDRAl". J. DROUET, P. AMOUCH, and M. Steinbuch 106
Protease Inhibitors in Human Synovial Fluids of Patients with J oint Diseases:
G. Feinstein!, G. SHTACHER, and R. MAAYAN. With 1 Figure 109
Proteinase Inhibitors of the Respiratory Tract: Studies on the Structural
Relationship between Acid-Stable Inhibitors Present in the Respiratory
Tract. Plasma and Urine: K. Hochstrasser, H.FEUTH, and K. HocH-
GESAND 111
* In the case of several authors bold-face type indicates the name of the author who
presented the paper at the conference.
1 Prevented from attending. paper not presented at the conference.
VIII Contents
Section II
Proteinases and Proteinase Inhibitors
in Fertilization
Biochemistry and Clinical Significance of the Trypsin-Like Proteinase
Acrosin from Boar and Human Spermatozoa: H.Fritz, W.D.SCHLELJ
NING, and W.-B. SCHILL. With 10 Figures . . . . . . . . . . . . . 118
Studies on the Purification and Characterization of Boar Acrosin:
K. L. Polakoski and W. L. WILLIAMS. With 9 Figures ........ 128
Proteinase Inhibitors and Proteinases of Human Semen: L. J. D. Zaneveld,
G. F. B. SCHUMACHER, P. F. TAUBER, and D. PROPPING. With 6 Figures 136
Characterization of Two Proteinase Inhibitors from Human Seminal Plasma
and Spermatozoa: H.Schiessler, M.ARNHOLD, and H.FRITZ. With 7
Figures ........................... 147
Isolation of Proteinase Inhibitors from Boar Sperm Acrosomes and Boar
Seminal Plasma and Effect on Fertilization: K. L. Polakoski and
W. L. WILLIAMS. With 8 Figures 156
Purification, Characterization, and Structural Studies of Proteinase Inhibitors
from Boar Seminal Plasma and Boar Spermatozoa: H. Tschesche,
S. KUPFER, O. LENGEL, R. KLAUSER, M. MEIER, and H. FRITZ. With 12
Figures ........................... 164
Proteinase Inhibitors in Human Cervical Mucus and Their in vitro Inter
actions with Human Acrosin: G. F. B. Schumacher and L. J. D. ZAN EVELD.
With 3 Figures . . . . . . . . . . . . . . . . . . . . . . . . 178
Section II I
Pancreatic Proteinase Inhibitors
Human Pancreatic Secretory Trypsin Inhibitor: L. J. Greene, D. E.RoARK,
and D.C.BARTELT. With 8 Figures . . . . . . . . . . . . . . . . 188
Isolation of Human Pancreatic Inhibitor and the Study of Its Interaction
with Mammalian and Human Proteases: G. Feinstein 2, R. HOFFSTEIN, and
M. SOKOLOVSKY. With 11 Figures ................ 199
Studies on Inhibition of the Two Human Trypsins: C. Figarella, G. A. NEGRI,
and O.GUY. With 13 Figures . . . . . . . . . . . . . . . . . . 213
Sequence of Cleavages in Temporary Inhibitors: Porcine Inhibitor Form II
and Bovine Isoinhibitors A and C: S. L. SCHNEIDER, L. ST ASIUK, and
M. Laskowski, Sr. With 9 Figures ................ 223
Temporary Inhibition - A Result of Consecutive Peptide Bond Hydrolysis:
H. Tschesche, G.REIDEL, and M.SCHNEIDER. With 3 Figures ..... 235
Measurement of the Bovine Pancreatic Trypsin Inhibitors by Radioim
munoassay: E. Fink and L. J. GREENE. With 6 Figures . . . . . . . . 243
Properties and Inhibition Spectrum of a New Human Pancreatic Protease:
P. A. MALLORY and J. Travis . . . . . . . . . . . . . . . . . . . 250
2 Prevented from attending, paper not presented at the conference.
Contents IX
Section IV
Characterization and Structure of New Inhibitors
Amino Acid Sequence of Snail Inhibitor K and Correlation of Structure and
Specificity: T. Dietl and H. TSCHESCHE. With 8 Figures ... . . . . 254
Novel Proteinase Inhibitors in Snake Venoms: Distribution, Isolation, and
Amino Acid Sequence: H. TAKAHASHI, S.IwANAGA, T.KITAGAWA,
Y.HOKAMA, and T.Suzuki. With 8 Figures ............ 265
Broad Specificity Inhibitors from Sea Anemones: G. Wunderer, K.KuMMER,
H. FRITZ, L. BERESS, and W. MACHLEIDT. With 4 Figures . . . . . . . 277
Discussion Remark: A Polyvalent Proteinase Inhibitor in the Earthworm
(Lumbricus terrestris): K. ILLCHMANN and E. WERLE: Presented by
H. FRITZ .......................... 282
Discussion Remark: Proteinase Isoinhibitors from Cuttle Fish (Loligo vul
garis): H. Tschesche and A. VON RUCKER. With 1 Figure . . . . . . . 284
The Partial Linear Sequence of Garden Bean Inhibitor II and Location of
the Protease Reactive Sites: K. A. Wilson and M. LASKOWSKI, Sr. With 2
Figures ........................... 286
Elastase Inhibitors as Impurities in Commercial Preparations of Soybean
Trypsin Inhibitor (Kunitz): J.Bieth and J.-C.FRECHIN. With 11 Figures 291
A New Alkaline Proteinase Inhibitor from Streptomyces albogriseolus:
T.IKENAKA. With 1 Figure . . . . . . . . . . . . . . . . . . . 305
Section V
Structure-Function Relationships
Replacements, Insertions, and Modifications of Amino Acid Residues in the
Reactive Site of Soybean Trypsin Inhibitor (Kunitz): D. Kowalski,
T.R.LEARY, R.E.McKEE, R.W.SEALOCK, D.WANG, and M.LASKOW-
SKI, Jr. With 8 Figures. . . . . . . . . . . . . . . . . . . . . . 311
Chemical Structure and Inhibitory Activities of Soybean Proteinase
Inhibitors: T.Ikenaka, S.ODANI, and T. KOIDE. With 23 Figures 325
Structure-Function Relationships in Lima Bean Protease Inhibitor:
F. C. Stevens, S. WUERZ, and J. KRAHN. With 8 Figures . . . . . . . 344
Structure-Activity Relationships of Several Trypsin and Chymotrypsin
Inhibitors from Legume Seeds: Y.BIRK. With 1 Figure ....... 355
Reactive Site Cleavage, Thermodynamic Control Resynthesis, and Properties
of Chemically Derivatized Trypsin-Kallikrein Inhibitors: H. TscHEscHE,
H.Jering, G.SCHORP, and T.DIETL. With 15 Figures ........ 362
The Interaction of Catalytically Inactive Proteolytic Enzymes with Protein
Inhibitors. Methyl-Chymotrypsin: D. S. RYAN and R. E. Feeney. With
3 Figures ........ ........ . . . . . 378
Discussion Remark: Inhibition of Anhydro-Trypsin and Anhydro-
Chymotrypsin: C. A. RYAN . . . . . . . . . . . . . 384
x
Contents
Section V I
Physical-Chemical Studies of Enzyme-Inhibitor Interactions
Kinetics and Thermodynamics of the Interaction of Proteinases with Protein
Inhibitors: W. R. Finkenstadt, M. A. HAMID, J. A. MATTIS, J. SCHRODE,
R. W. SEALOCK, D. WANG, and M. LASKowsKI,Jr. With 16 Figures 389
Kinetic Studies of the Binding of Bovine Basic Pancreatic Trypsin Inhibitor
to a-Chymotrypsin: J. Engel, U. QUAST, H. HEUMANN, G. KRAUSE, and
E.STEFFEN. With 5 Figures ................... 412
The Mechanism of Association of Trypsin (or Chymotrypsin) with the
Pancreatic Trypsin Inhibitors (Kunitz and Kazal), Kinetics and Thermo
dynamics of the Interaction: M.Lazdunski, J.-P. VINCENT, H. SCHWEITZ,
M.PERoN-RENNER, and J.PUDLES. With 7 Figures ......... 420
Kinetic Studies on the Interactions between Native, Acetylated and Suc
cinylated Trypsin and Natural Proteinase Inhibitors: N. Levilliers,
M.PERoN-RENNER, and J.PUDLES. With 5 Figures ......... 432
The Use of Fluorescence Techniques in the Study of the Interaction of the
Basic Trypsin Inhibitor of Bovine Pancreas, Selectively Labelled at Lysine
15, with Chymotrypsin and Trypsin: Y.ELKANA (presented by M.RIGBI).
With 5 Figures ........................ 445
Raman Studies of Native and Partially (Cys 14-Cys 38) Reduced Basic
Pancreatic Trypsin Inhibitor: H. BRuNNER and M. HOLZ (presented by
H.JERING). With 2 Figures ... . . . . . . . . . . . . . . . . 454
A Study on the Interaction of Trasylol with Trypsin and Kallikrein by
Circular Dichroism: H.RoSENKRANZ. With 4 Figures . . . . . . . . 458
Some Kinetic Consequences of the Tight Binding of Protein-Proteinase
Inhibitors to Proteolytic Enzymes and Their Application to the Deter
mination of Dissociation Constants: J. BIETH. With 2 Figures . . . . 463
Section VII
Molecular Aspects of Enzyme-Inhibitor Interactions
Stereochemistry of Substrate Binding and Hydrolysis in the Trypsin
Family of Enzymes: D.M.BLOw. With 9 Figures . . . . . . 473
The Model of the Basic Pancreatic Trypsin Inhibitor Refined at 1.5 A
Resolution: J.Deisenhofer and W.STEIGEMANN. With 11 Figures 484
Structure of the Complex Formed by Bovine Trypsin and Bovine Pancreatic
Trypsin Inhibitor, Refinement of the Crystal Structure Analysis: R.Hu
ber, D.KuKLA, W.STEIGEMANN, J.DEISENHOFER, and A. JONES. With 9
Figures ........................... 497
The Mode of Action of Soybean Trypsin Inhibitor as Revealed by Crystal
Structure Analysis of the Complex with Porcine Trypsin: J.JANIN,
R.M.Sweet, and D.M.BLOw. With 6 Figures ........... 513
Contents XI
Section VIII
Organic Chemical Models and Low-Molecular-Weight Inhibitors
Organic Chemical Models for Proteinase Inhibitors: E. T. KAISER 523
Progress in Designing Small Inhibitors Which Discriminate among Trypsin-
Like Enzymes: E.SHAW. With 8 Figures ..... . . . . . . . . 531
Homoarginine Oligomers as Trypsin and Chymotrypsin Substrates and In
hibitors: M.Rigbi, Y.ELKANA, N.SEGAL, D.KuGER, and L.SCHWARTZ.
With 6 Figures ........................ 541
Proteinase Inhibitors of Microbial Origin. A Review: W. WINGENDER. With
5 Figures .......................... 548
Discussion Remark: Low Molecular Weight Carboxydipeptidase Inhibitors
from Bothropsjararaca Venom: L.J.GREENE ............ 560
Section IX
Inhibition of Non-Serine Prot eases
Carboxypeptidase Inhibitor from Potatoes. Properties and Amino Acid
Sequence: C.A.Ryan, G.M.HASS, R.W.KuHN, and H.NEURATH. With
7 Figures . . . . . . . . . . . . . . . . . . . . . . . . . . . 565
Inhibitors of Lysosomal Proteinases: A. J. BARRETT. With 3 Figures . . . . 574
Inhibition of Insulin Specific Protease by Proteinase Inhibitors from the
Snail (Helix pomatia): J.S.BRUSH and H.Tschesche. With 3 Figures 581
An Insulin Degrading Proteinase from Human Erythrocytes and Its In
hibition by Proteinase Inhibitors: H. Tschesche, T. DIETL, H. J. KOLB,
and E.STANDL. With 6 Figures ................. 586
Section X
Evolution of Specificity of Proteinase Inhibitors
Evolution of Specificity of Protein Proteinase Inhibitors: M.Laskowski,Jr.,
I. KATO, T. R. LEARY, J. SCHRODE, and R. W. SEALOCK. With 10 Figures 597
Section XI
Proteinases and Inhibitors in Biological Systems
Proteases and Protease Inhibitors in Neoplasia: H. P. SCHNEBLI . . . . . 615
Secretion of Enzymes by Neoplastic Cells and Macrophages: E. REICH.
With 4 Figures ........................ 621
Plasminogen Activators of Normal and Transformed Cells: A.R.Goldberg
and S.G.LAZAROWITZ. With 5 Figures .............. 631
XII Contents
Proteinase Inhibitors from Ascaris lumbricoides: Properties and Their
Physiological Role: R.J.Peanasky, G.M.ABU-ERREISH, C.R.GAusH,
G.A.HOMANDBERG, D.O'HEERoN, R.K.LINKENHEIL, U.KUCICH, and
D.R.BABIN. With 18 Figures .................. 649
W ound-Induced Accumulation of Proteinase Inhibitor I in Tomato Leaves:
C.A.RYAN. With 2 Figures .............. . . . . . 667
Section XII
General Discussion Remark, Resume, Concluding Remarks
General Discussion Remark: Energetics of Inhibitor Binding. D. M. BLOW 677
Resume: M.LASKOWSKI,Jr. ............... 679
Concluding Remarks: H. TscHEscHE, L. J. GREENE, and H. FRITZ 685
Author Index 687
Subject Index 735
