Table Of Content2
VOLUME
Bacterial
Immunoglobulin-
Binding Proteins
Applications in
Immunotechnology
Edited by
Michael D. P. Boyle
Department of Microbiology
Medical College of Ohio
Toledo, Ohio
Academic Press, Inc.
Harcourt Brace Jovanovich, Publishers
San Diego New York Boston
London Sydney Tokyo Toronto
NOTE: The University of Florida holds patents for the isolation and use of type lib and
type III Fc-binding proteins. These patents have been licensed to Gator
Microbiologicals, Inc., a company in which Drs. Boyle and Faulmann have a financial
interest. Although we do not believe that this has influenced our interpretation of any of
the data presented in this volume, we believe that the reader should be aware of this
interest.
This book is printed on acid-free paper. @
Copyright © 1990 by Academic Press, Inc.
All Rights Reserved.
No part of this publication may be reproduced or transmitted in any form or
by any means, electronic or mechanical, including photocopy, recording, or
any information storage and retrieval system, without permission in writing
from the publisher.
Academic Press, Inc.
San Diego, California 92101
United Kingdom Edition published by
Academic Press Limited
24-28 Oval Road, London NW1 7DX
Library of Congress Cataloging-in-Publication Data
(Revised for vol. 2)
Bacterial immunoglobulin-binding proteins.
Includes bibliographical references.
Contents: v. 1. Microbiology, chemistry, and
biology ~ v. 2. Applications in immunotechnology.
1. Bacterial immunoglobulin-binding proteins.
1. Boyle, Michael D. P. [DNLM: 1. Bacterial Proteins.
2. Carrier Proteins. 3. Immunoglobulins. 4. Receptors,
Immunologie. QW 601 B131]
QR92.I4B33 1990 616'.014 89-6995
ISBN 0-12-123011-2 (v. 1 : alk. paper)
ISBN 0-12-123012-0 (v. 2 : alk. paper)
Printed in the United States of America
90 91 92 93 9 8 7 6 5 4 3 2 1
Contributors
I
Numbers in parentheses indicate the pages on which the author's contributions begin.
Bo Akerström (91), Department of Physiological Chemistry, University
of Lund, S-223 62 Lund, Sweden
Patrick Alexander1 (417), Genex Corporation, Gaithersburg, Maryland
20877
Elia M. Ayoub (161), Department of Pediatrics, University of Florida,
College of Medicine, Gainesville, Florida 32610
Douglas J. Barrett (393), Department of Pediatrics, University of Flor-
ida, College of Medicine, Gainesville, Florida 32610
Lars Björck (91), Department of Medical Microbiology, University of
Lund, S-223 62 Lund, Sweden
Michael D. P. Boyle (1, 23, 49, 71, 105, 145, 161, 181, 273, 291, 369, 405,
425), Department of Microbiology, Medical College of Ohio, Toledo,
Ohio 43699
L. Jeannine Brady (161), Department of Oral Biology, University of
Florida, College of Dentistry, Gainesville, Florida 32610
Colleen Chun (161), Department of Pediatrics, University of Florida,
College of Medicine, Gainesville, Florida 32610
Sylvia E. Coleman (217), Department of Microbiology and Cell Science,
University of Florida, Gainesville, Florida 32610
Stephen R. Fahnestock2 (417), Genex Corporation, Gaithersburg, Mary-
land 20877
1 Present address: Center for Advanced Research in Biotechnology, Gaithersburg, Maryland
20877.
2 Present address: National Institute of General Medical Sciences, Bethesda, Maryland.
xv
xvi Contributors
Ervin L. Faulmann (49, 71, 125, 249, 273), Department of Microbiology,
Medical College of Ohio, Toledo, Ohio 43699
Adrian P. Gee (181, 405), Baxter Health Care Corporation, Fenwal
Division, Santa Ana, California 92705
Hector Juarez-Salinas (341), Chromatography Business Unit, Bio-Rad
Laboratories, Richmond, California 94806
Michael J. P. Lawman (181, 291, 405), Department of Immunology and
Medical Microbiology, University of Florida, College of Medicine,
Gainesville, Florida 32610
Patricia D. Lawman (405), Department of Oral Biology, College of Medi-
cine, and Department of Immunology/Medical Microbiology, College
of Dentistry, University of Florida, Gainesville, Florida 32610
Lawrence J. Mclntyre (205), Vector Laboratories, Inc., Burlingame,
California 94010
Corey Musselman (161), Department of Immunology and Medical Micro-
biology, University of Florida, College of Medicine, Gainesville, Flor-
ida 32610
Ronald A. Otten (49, 425), Department of Microbiology, Medical Col-
lege of Ohio, Toledo, Ohio 43699
KathleenJ.Reis3(23,49,71,105,145,291), Department of Large Animal
Clinical Sciences, College of Veterinary Medicine, University of Flor-
ida, Gainesville, Florida 32610
Larry Schwartz (309), Technical Service, Pharmacia LKB Biotechnology
Inc., Piscataway, New Jersey 08854
Susan M. Scott (341), Chromatography Business Unit, Bio-Rad Labora-
tories, Richmond, California 94806
Edward J. Siden (301), Division of Clinical Immunology, Department of
Medicine, Mount Sinai School of Medicine, New York, New York
10029
Stephen N. Sisson (197), Cascade Immunology Corporation, Springfield,
Oregon 97478
Barbara Webb Walker (355), Genex Corporation, Gaithersburg, Mary-
land 20877
3 Present address: Genex Corporation, 16020 Industrial Drive, Gaithersburg, Maryland
20877.
Preface
Volume 1 brought together in a single book the current state of knowledge
of bacterial immunoglobulin-binding proteins. In this volume the focus is
on practical approaches to isolation, characterization, and use of these
binding proteins. The majority of these studies involve the type I Fc-
binding protein (staphylococcal protein A) and the type III Fc-binding
protein (streptococcal protein G). These proteins represent the prototypes
of a larger family of functionally related bacterial immunoglobulin-binding
proteins. The applications described in this volume for the prototype
molecules should be adaptable to any new selective binding proteins that
become available. An attempt has been made by all the contributors to
provide sufficient information to enable any investigator to use bacterial
IgG-binding proteins for the specific purpose described without having to
consult any secondary references. In addition, the limitations of individual
techniques and practical problems experienced by investigators have been
stressed. Although it is noted in many of the chapters in this volume, I
would like to remind the reader that the species and subclass reactivity of
bacterial immunoglobulin-binding proteins is not absolute. In particular,
with monoclonal antibodies it is not always possible to predict reactivity
just by knowing isotype and subclass.
Finally, I would like to thank the following people: everyone in my
laboratory who has contributed to the techniques and methods described;
my wife, Carla, and my children, Kieron and Sarah, for their long suffering
during the compilation; and my secretary, Shirley Doherty, who battled
my handwriting to get this volume to the publisher.
Michael D. P. Boyle
xvii
1
CHAPTER
I
Introduction to bacterial
immunoglobulin-binding
proteins
Michael D. P. Boyle
I. Introduction
In the early part of the twentieth century, Landsteiner observed that red
cells from one individual could be agglutinated by serum from certain other
individuals. These findings led to the understanding of blood group anti-
gens and the birth of serological tests for identifying antigens on cells. This
hemagglutination assay is still widely used today as the basis for rapid
screening and typing assays in blood banks. With the recognition that
specific antibodies could neutralize toxins (von Behring and Kitasato,
1890) or cause selective precipitation of soluble antigens (Heidelberger
and Kendall, 1932), semiqualitative methods for identifying antigens and
antibodies were developed. More recently, the sensitivity of antigen or
antibody detection has been increased with the development of radioim-
munoassays (RIA) (Yallow and Benson, 1960) and enzyme-linked immu-
nosorbent assays (ELISA) (Engvall and Perlmann, 1971, 1972). These
techniques now enable the determination of absolute levels of antigens or
antibodies in the nanomolar range or below and have found broad applica-
tions for clinical diagnostic procedures and in forensic medicine.
In 1975, Köhler and Milstein described a method for fusing myeloma
cells with spleen cells from immunized mice that enabled the selection of a
hybrid cell line that produced large quantities of a single antibody. This
monoclonal antibody technology has the potential to allow antibodies
specific for any desired epitope to be produced. This technology has had a
profound impact on immunotechnology and has enabled the development
of many new antibody-based detection systems. In particular, the produc-
Bacterial Immunoglobulin-Binding Proteins, Volume 2
Copyright © 1990 by Academic Press, Inc. All rights of reproduction in any form reserved. 1
2 Michael D. P. Boyle
tion of monoclonal antibodies to T cell surface markers (Reinherz et al.,
1979) coupled with the development of the fluorescence-activated cell
sorter has revolutionized many aspects of clinical immunology (for review
see Shapiro, 1985; Ault, 1986; Jackson and Warner, 1986). The ability to
relate the phenotypes of lymphocytes to their functions, based on their
reactivities with specific monoclonal antibodies that recognize surface
glycoproteins or cluster determinants, has enabled the clinical immunolo-
gist to identify and characterize a variety of acquired and inherited immu-
nodeficiency disorders (for review see Giorgi, 1986). In particular, the
inversion of the ratio of CD4 to CD8 T lymphocytes in patients with
acquired immunodeficiency syndrome (AIDS) is now characteristic for
that disease (Centers for Disease Control (CDC), 1982; Rosenberg and
Fauci, 1989).
Over the past decade more sophisticated methods of quantifying and
characterizing antigens in complex mixtures and on cell surfaces have
been developed. All of these methods require the ability to produce and
isolate monospecific antibodies (monoclonal or polyclonal) and to detect
these antibodies when complexed with their specific antigens. Proteins
can be separated on sodium dodecyl sulfate (SDS)-polyacrylamide gels by
virtue of their size. The proteins can then be transferred to nitrocellulose
and probed with specific antibody to identify individual antigens, allowing
them to be both quantified and characterized for size heterogeneity (Tow-
bin et al., 1979). These Western blotting approaches are now finding wide
use both for research applications and for immunodiagnostic purposes.
With the increased interest in immunotechnology for analytic methods,
quantitative determination of antigens, measurement of antibodies, and
monitoring and isolation of specific immunoglobulins, much interest has
focused on reagents that facilitate these immunological techniques. Bacte-
rial immunoglobulin-binding proteins represent such a family of valuable
immunological reagents (Langone, 1978, 1982b; Langone et al., 1977,
1979; Boyle, 1984; Boyle and Reis, 1987; Forsgren et al., 1983; Richman,
1983;Duboid-Dalcqera/., 1977; Change/., 1984;Goding, 1978; Gee and
Langone, 1981). The purpose of this volume is to provide a practical guide
to the uses of bacterial immunoglobulin-binding proteins, in particular
those that bind selectively to constant regions of immunoglobulin mole-
cules, without interfering with the ability of the antibody to bind to its
specific antigen. The selection and isolation of these unique groups
of bacterial proteins and their application both to the isolation of anti-
body molecules and to the detection of antigen-antibody complexes
will be described in detail. In this chapter, a brief survey of bacterial
immunoglobulin-binding proteins and their reactivities is presented. For a
more comprehensive treatment, see Volume 1 of this series.
Chapter 1. Introduction 3
IL The Distribution and Functional Reactivity of Bacterial
IgG Fc-Binding Proteins
Bacterial Fc-binding proteins have been found on the surface of a variety
of streptococci and staphylococci, and more recently on other organisms
(Tables 1 and 2). Bacteria expressing IgG-binding proteins have been
detected by a variety of methods, including their ability to agglutinate red
cells that have been sensitized with subagglutinating doses of specific
antibody and their ability to bind labeled IgG via regions of the immuno-
globulin molecule not involved in specific antigen recognition (Chapter 2).
Early studies using different species and subclasses of IgG, have demon-
strated five distinct patterns of IgG binding to intact bacteria, which led
Myhre and Kronvall (1981) to propose a functional classification for bacte-
rial IgG Fc-binding proteins (Figure 1). More recently, we have identified
a bacterial isolate that demonstrates a sixth profile of binding to IgG
from different mammalian species and we have designated this reactiv-
ity as type VI (Figure 1). All of these different functional types of
immunoglobulin-binding proteins appear to be mediated by antigenically
TABLE 1
Bacterial IgG Fc-Binding Proteins0
Type Bacterial species References
Type I Staphylococcus aureus (protein A) Forsgren and Sjöquist (1966);
Sjöquist et al. (1972);
Langone (1982a)
Type II Group A streptococci Havlicek(1978);
Gvubb et al. (1982);
Yarnall and Boyle (1986a-c)
Type III Streptococcus equisimilus (group C) Reis et al. (1984a,b)
Streptococcus dysgalactiae (group C) Björck and Kronvall (1984)
Human group G streptococci
Human group G streptococci
Type IV Bovine ß-hemolytic group G Myhre et al. (1979);
streptococci Reis?/A/. (1990)
Type V Streptococcus zooepidemicus Myhre and Kronvall (1980);
(group C) Yarnall and Widders (1990)
Type VI Streptococcus zooepidemicus Reisetal. (1988)
(group C)#S212
a Classification originally proposed by Myhre and Kronvall (1981) with the addition of type VI
proposed by Reis et al. (1988).
4 Michael D. P. Boyle
TABLE 2
Types of Bacteria that Bind Immunoglobulin in a Nonimmune Fashion
Bacteria IgG IgM IgA IgD IgE References
Streptococcus, Group B + Russell-Jones et al. (1984);
Brady and Boyle (1989)
+ + Jürgens et al. (1987)
Branhamella catarrhalis + Forsgren and Grubb ( 1979)
Clostridium perfringens + + Lindahl and Kronvall (1988)
Taylorella equigenitalis + + Widders et al. (1985)
Brucella abortus ( + )" Nielsen É·/a/. (1981)
Coprococcus comes + Van der Merwe and
Stegeman (1988)
Peptococcus magnus + + + + Myhre and Erntell (1985)
Björck(1988)
Haemophilus somnus + Yarnalle/a/. (1988)
a Reactivity limited to a subgroup of bovine IgM antibodies.
distinct proteins (Boyle and Reis, 1990) and are associated with distinct
bacterial species (Table 1).
In addition to the six types of bacterial IgG-binding proteins, immuno-
globulin-binding proteins for other immunoglobulin isotypes have been
identified on other bacteria (Table 2). The properties of these various
types of bacterial immunoglobulin-binding proteins are summarized in
Section III.
III. Bacterial IgG-Binding Proteins
The chemistry, microbiology, and functional activity of these molecules
have been reviewed in a comprehensive manner in Volume 1 of this series.
In the next section of this chapter, the properties of certain of these
molecules are summarized briefly with respect to their practical values in
immunotechnology.
A. Type I
The type I Fc-binding protein is found on the majority of Staphylococ-
cus aureus strains and more frequently designated as staphylococcal
Chapter 1. Introduction 5
Type I Type II Type III Type IV Type V Type VI
Human
Rabbit
Pig
Mouse ·
Rat
Cow ·
Goat
Sheep
Cat
Dog ^B O · nt
FIGURE 1
Profile of species reactivities of different bacterial IgG-binding protein types. Larger clots
represent greater binding activity; o, no reactivity, nt, not tested. Note these reactivities
have been determined using polyclonal IgG preparations. Occasional differences have
been noted in the reactivity of samples from individual animals. (Reproduced with
permission from Boyle and Reis 1990.)
protein A. This protein has been extensively characterized (for review see
Langone, 1982a; Boyle, 1990) and the gene coding for it has been cloned
and sequenced (Duggleby and Jones, 1983; Uhlén et al., 1984; Löfdahl et
ai, 1983; Guss et aL, 1990). Protein A's binding activity to a variety of
different species, classes, and subclasses has been well documented (Ta-
bles 3 and 4).
