9 780854 046980

ISBN 978-0-85404-698-0

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Molecular Aspects of Innate and Adaptive Immunity

Edited by Kenneth B M Reid and Robert B Sim

Reid & SimM

olecular Aspects of

Innate and Adaptive Im

munity

This book provides a survey of topics, in the area of innate and adaptive immunity, which have been researched within the MRC Immunochemistry Unit, at Oxford University, over a period of forty years. The topics include:

Antibody structure - for which the first Director of the Immunochemistry Unit, Professor RR Porter, was awarded a Nobel prize in 1972

The characterization of membrane proteins on lymphoid cells – leading to the concept of these molecules belonging to an immunoglobulin super family

The proteins of the human serum complement system – one of the body’s major defences against microbial infection

The human cell surface integrins and the hyaluronan-binding proteins, which are involved in regulation of inflammation at cell surfaces and within the extracellular matrix

The family of collectin molecules – containing distinct globular carbohydrate-binding domains linked to collagen-like regions – which play important roles in innate immunity in the lungs and bloodstream by immediate recognition and clearance of microbial pathogens

Each chapter in the book gives a brief historical background to a topic followed by a survey of recent advances in the field, all written by internationally recognised renowned experts. The theme running through the chapters is that of protein structure-function relationships including, amongst other things, a description of the mechanism of how the reactive thiol ester bonds, found in the complement system components C3 and C4, are activated to allow the covalent binding of these proteins to suitable targets on microbes.

Molecular Aspects of Innate and Adaptive Immunity is aimed primarily at established senior research scientists, postdoctoral research scientists and PhD students who have an interest in proteins of the immune system. The wide range of topics in immunity systems, while staying broadly within innate and adaptive immunity, will also appeal to a wider audience.

Front cover image courtesy of H.R. Knobel, W. Villiger and H. Isliker, Chemical analysis and electron microscopy studies of human C1q prepared by different methods, Eur. J. Immunol., 1975, 5, 78-82. © Wiley-VCH Verlag GmbH & Co. KGaA. Reproduced with permission.

Molecular Aspects of Innate and Adaptive Immunity

Dedication

This book is dedicated to all former members of the Medical Research CouncilImmunochemistry Unit, 1967–2008, and especially in remembrance of the staffmembers Rod Porter, Rachel Fruchter, Lawrence Mole, Alan Williams, TonyGascoyne, Audrey Richards, and visiting scientists Marion Koshland, MettaStrand, Walter Palm and Gordon Ross.

Molecular Aspects of Innateand Adaptive Immunity

Edited by

Kenneth BM Reid and Robert B Sim

MRC Immunochemistry Unit, Department of Biochemistry, University ofOxford, Oxford, UK

ISBN: 978-0-85404-698-0

A catalogue record for this book is available from the British Library

rRoyal Society of Chemistry, 2008

All rights reserved

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Preface

This volume provides a survey of topics in the area of innate and adaptiveimmunity, which have been researched within the MRC ImmunochemistryUnit, Oxford University, over a period of 40 years. The Unit was formed in1967, with Professor R. R. Porter (who was awarded a Nobel Prize in 1972 forhis research on antibody structure) as its first Director, with Professor K. B. M.Reid being Director from 1986 onwards.The extension of the research on antibody structure into a study of one of the

body’s major defence systems, the complement system (which can be triggeredby many stimuli including antibody recognition of invading microbes) led tothe molecular characterization of many of the proteins of this important cas-cade system. Particular attention was paid to the structure–function relation-ships and genetics of the early components of the system, especiallycomponents C1q, C1r, C1s, C2, factor B, factor D, the allotypes of C4, thecovalent binding properties of activated C3 and C4, and also the proteins thatregulate complement activation – such as C1inhibitor, factors H and I, C4b-binding protein and properdin. Studies of complement receptor specificity andstructure were also undertaken, as well as a range of studies on complementevolution and phylogeny.The domain organization and functions of human leukocyte integrins (two of

which are complement receptors), particularly with respect to heterodimerformation and activation of integrins, analysis of their deficiency [leukocyteadhesion deficiency (LAD)] and the involvement of these molecules in therecognition and resolution of inflammation, was another long-term researchtopic.The study of the molecular genetics of complement component C4 led on to

the analysis of a range of disease-susceptibility genes within the human majorhistocompatibility complex (MHC) Class III region, and of associated novelgenes within the human MHC. These included heat shock protein (HSP) 70,tumour necrosis factor (TNF) alpha and cytochrome (Cyt) P450 loci and theirassociation with immune-related diseases, a topic that has been a majorresearch interest within the Unit.

Molecular Aspects of Innate and Adaptive Immunity

Edited By Kenneth BM Reid and Robert B Sim

r Royal Society of Chemistry, 2008

Published by the Royal Society of Chemistry, www.rsc.org

v

The characterization of membrane proteins on leukocytes was initiated in theearly 1970s, and led to the concept that many of these molecules belong to animmunoglobulin super family. This research was initiated by Alan F. Williamsin the Immunochemistry Unit and continued, from 1978 onwards, in the MRCCellular Immunology Unit.In more recent years, structural and functional studies of hyaluronan (HA)-

binding proteins and receptors, involved in extracellular matrix remodellingand leukocyte migration during inflammation, formed a major research focusin the Unit. HA is a high molecular weight polysaccharide found in tissues of allvertebrates. The interaction of HA-binding proteins with HA has a central rolein the formation and stability of extracellular matrix (e.g. in cartilage) and isalso important in immune-cell trafficking. The research has concentrated onTSG-6 (the protein product of TNF-stimulated gene-6), an HA-binding proteinwhich is only expressed in adult tissues during inflammation, and the ubiqui-tous HA-receptor CD44, which is involved in matrix assembly and leukocytemigration.Another major interest within the Immunochemistry Unit has involved the

family of collectin molecules – which contain distinct globular carbohydrate-binding domains linked to collagen-like regions – and play important roles ininnate immunity in the lungs and bloodstream by immediate recognition andclearance of microbial pathogens. This was most recently extended to include theficolins, which have structural and functional similarities to the collectins. Theserum collectin mannan-binding lectin (MBL) and ficolins recognize arrays ofneutral sugars or acetylated compounds on microorganisms as being foreignand, after binding to them, stimulate activation of MBL-associated serineproteases (MASPs), which result in activation of the serum complement system –thus recruiting one of the major defence systems in the body. Collectins found inlung surfactant (surfactant proteins A and D (SP-A and SP-D)) also recognizeand clear microorganisms, but without involvement of complement activation.The collectins and ficolins, in addition to their interactions with microorganisms,can resolve inflammation by binding to and promoting clearance of apoptoticcells and DNA – and by modulating the manner in which allergens are processedby dendritic cells. Recombinant forms of the collectins and ficolins are con-sidered as candidates for new therapeutics for the treatment of infection andinflammation.Each chapter in the volume gives a brief historical background to a topic,

with some emphasis on work carried out within the Immunochemistry Unit,and then provides a survey of recent advances in the field. The main themerunning through most of the chapters is that of protein structure–functionrelationships – good examples being the descriptions of quaternary structuresof large oligomeric proteins, of factor H and C1q binding to specific ligands, orof the chemistry of the mechanism of catalysis of the covalent binding ofactivated C3 and C4 proteins to nucleophilic groups on microbial surfaces.

Kenneth B. M. ReidRobert B. Sim

vi Preface

Contents

Section 1 Antibodies

Chapter 1 R. R. Porter and the Structure of Antibodies

Lisa A. Steiner and Julian B. Fleischman

1.1 Introduction 31.2 The Papain Fragments 41.3 The Four-chain Model 71.4 V and C Regions of Immunoglobulin Chains 8References 9

Chapter 2 Chemical Engineering of Therapeutic Antibodies

George T. Stevenson and Weng Leong

2.1 Introduction 112.1.1 Structure of IgG 112.1.2 Availability of SH Groups 132.1.3 Enzymic Dissections 13

2.2 Useful Chemistry of SH Groups and SS Bonds 142.2.1 SS Interchange 152.2.2 Alkylation of SH Groups by Maleimides 17

2.3 Engineering IgG Modules 182.3.1 Attaching the PDM Linker to Fcg 192.3.2 Attaching the PDM Linker to Fab0g 202.3.3 Examples of Therapeutic Constructs 21

Acknowledgements 23References 23

Molecular Aspects of Innate and Adaptive Immunity

Edited By Kenneth BM Reid and Robert B Sim

r Royal Society of Chemistry, 2008

Published by the Royal Society of Chemistry, www.rsc.org

vii

Section 2 The Complement System

Chapter 3 The Evolution of Complement Systems

Alister W. Dodds

3.1 Phylogeny in the MRC Immunochemistry Unit 273.2 Evolution of Immune Systems 283.3 Evolution of the Complement System 30

3.3.1 The Complement Pathways 303.3.2 Loss and Gain of Complement Components

in Some Evolutionary Lines 313.4 Phylogeny of the Complement System 33

3.4.1 Recognition Molecules; Ficolins, MBL andC1q 34

3.4.2 Initiation Enzymes; Factor D, MASPs, C1rand C1s 37

3.4.3 Thioester Proteins: C3, C4, C5, a2M, CD109and Insect Thioester Proteins 39

3.4.4 Amplification Enzymes; C2 and Factor B 403.4.5 Lytic Proteins; C6, C7, C8 and C9 413.4.6 Control Proteins and Receptors 413.4.7 Possible Origins of Complement 42

3.5 Summary 43Acknowledgements 43References 44

Chapter 4 Structure and Function of the C1 Complex: A Historical

Perspective

Gerard J. Arlaud

4.1 From the Antibody to the C1 Complex of

Complement 494.2 Biochemistry of C1q and Elucidation of its Primary

Structure 504.3 Biochemistry of C1r and C1s and the C1 Activation

Mechanism 534.3.1 Biochemistry of C1r and C1s 534.3.2 Insights into the C1 Activation Mechanism 534.3.3 The Primary Structure of C1r 54

4.4 From the Domain Structure of C1r and C1s

to a Model of C1 Architecture 554.4.1 The N-terminal Interaction Domains 554.4.2 The C-terminal Catalytic Domains 564.4.3 A Low-resolution C1 Model 57

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4.5 The Era of Structural Biology 574.5.1 A Three-dimensional C1 Model 574.5.2 C1r Activation and the Triggering Signal 584.5.3 C1q: Binding Versatility Arises fromModularity 60

4.6 Conclusions and Perspectives 61References 61

Chapter 5 Complement Components C3 and C4

S. K. Alex Law

5.1 The Road to Oxford 655.2 The Autolytic Cleavage Reaction 675.3 Red Wine and the Isotypes of C4 695.4 LOO-3, the Anti-C4Monoclonal Antibody that Only

Worked for Us 705.5 How to Catch Up from Three Years Behind 725.6 C4K and C4Y 745.7 The Best Beer that I had Ever Tasted 765.8 Silence is Golden, but could be Painful 765.9 A Bet that I both Won, and Lost 77

5.10 Final Comments 79Acknowledgements 80References 81

Chapter 6 Complement Control Proteins and Receptors:

From FH to CR4

Robert B. Sim, Beryl E. Moffatt, Jacqueline M. Shaw and

Janez Ferluga

6.1 Introduction 846.2 Factor H 856.3 Factor I 936.4 C4b-Binding Protein and Properdin 956.5 Complement Receptors 966.6 Complement Research into its Third Century 98Acknowledgements 98References 99

Chapter 7 Biology and Genetics of Complement C4

Michael C. Carroll

7.1 Introduction 1057.2 Early Days in Oxford 1057.3 Linkage of Innate and Adaptive Immunity 1087.4 C4 and Autoimmunity 109

ixContents

7.5 Anti-RNA Mouse Model 1117.6 Models for C4 in B Cell Tolerance 1137.7 Summary 114References 114

Section 3 Collectins and Ficolins in Innate Immunity

Chapter 8 The Structure of Mannan-binding Lectin and its

Functional Relevance

Jinhua Lu and Steffen Thiel

8.1 Introduction 1218.2 MBL was Purified as a ‘By-product’ 1228.3 MBL Structure, Genetics and Heterogeneity in Size 1238.4 Concluding Remarks 126References 127

Chapter 9 Personal Accounts of the Discovery of MASP-2 and its Role

in the MBL Pathway of Complement Activation

Thomas Vorup-Jensen and Jens Chr. Jensenius

9.1 Preamble 1299.2 The Oxonian Connection 1309.3 MASP and the Serine Proteases of the

Complement System 1319.4 Gene Characterization in the Pre-genomic Age:

Discovering MASP-2 1349.5 Beyond Finding MASP-2 1409.6 MASP-3: One Gene, Two Enzymes 1429.7 End Note 143References 144

Chapter 10 The Structure and Function of Ficolins, MBLs and MASPs

Russell Wallis, Anders Krarup and Umakhanth Venkatraman

Girija

10.1 Introduction 14710.2 MBL and Ficolins in the Innate Immune System 14810.3 Genetics of MBLs and Ficolins 148

10.3.1 Genetics and Tissue Distributions of MBLs 14810.3.2 Genetics and Tissue Distributions of

Ficolins 15110.4 Structural Organization of MBLs and Ficolins 15210.5 Target Recognition by MBLs and Ficolins 155

10.5.1 Sugar Recognition by MBLs 15510.5.2 Ligand binding by Ficolins 155

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10.6 Interactions between MBLs, Ficolins and

MASPs 15710.7 Mechanism of Complement Activation in the Lectin

Pathway 16110.8 Conclusions 161References 162

Chapter 11 Surfactant Protein D and Glycoprotein 340

Jens Madsen and Uffe Holmskov

11.1 Introduction 16711.2 Domain Organization and Expression 16811.3 Gp-340, Agglutinin, DMBT1 and Immune Functions 16911.4 DMBT1gp-340 and its Interaction with SP-D 17011.5 DMBT1 and its Interaction with Other Host

Molecules 17111.6 Conclusion 172References 173

Section 4 Cell Surface Proteins – Immunoglobulin Superfamily and

Integrins

Chapter 12 Leukocyte Surface Proteins – Purification and

Characterization

A. Neil Barclay

12.1 Background 18112.1.1 Quantitation of Cell Surface Proteins 18112.1.2 Was there a Receptor on T Cells that

Incorporated Ig Variable Regions? 18312.1.3 Assays that Worked in Detergent 18412.1.4 Thy-1 – The Pioneer for Characterizing

a Lymphocyte Membrane Protein 18412.1.5 Solubilization of Surface Proteins 18412.1.6 Purification Using Lectin Affinity Columns 18512.1.7 Antibody Affinity Columns 18512.1.8 Biochemical Analysis of Thy-1 18612.1.9 Amino Acid Sequence Analysis 186

12.1.10 Monoclonal Antibodies to Recognize NewCell Surface Proteins 187

12.1.11 The Immunoglobulin Superfamily Concept 18712.1.12 The Follow-up 188

12.2 The Legacy 191Acknowledgements 191References 191

xiContents

Chapter 13 Cell Surface Integrins

Suet-Mien Tan and S. K. Alex Law

13.1 Introduction 19513.2 From Complement Proteins to Integrins 19513.3 Integrins as Modular Proteins 19813.4 The Flow of Conformational Changes in Integrin

During its Activation 20213.5 Concluding Remarks 208Acknowledgements 208References 208

Section 5 Immunogenetics and Major Histocompatibility Complex

Class III Analysis

Chapter 14 Molecular Genetics of the Major Histocompatibility

Complex Class III Region

R. Duncan Campbell, Wendy Thomson and Bernard Morley

14.1 Foreword by Duncan Campbell 21914.2 Physical Mapping of the Major Histocompatibility

Complex 22014.3 Sequence Analysis of the MHC and Annotation

of the Genes 22214.4 Genetic Analysis of the Complement Genes 224

14.4.1 Polymorphism of C4 22414.4.2 Deficiency of C4 22514.4.3 Polymorphism and Deficiency of C2 22614.4.4 Polymorphism and Deficiency of Factor B 226

14.5 Disease Association Studies 22714.5.1 Candidate Gene Studies 22714.5.2 Multiple Marker Studies 229

14.6 Conclusions 229References 230

Chapter 15 Functional Characterization of Major Histocompatibility

Complex Class III Region Genes

R. Duncan Campbell, Caroline M. Milner and

Begona Aguado

15.1 Introduction 23815.2 Genes that Encode Cell Surface Receptors 239

15.2.1 G6B 23915.2.2 G6F 244

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15.2.3 Lymphocyte Antigen 6 SuperfamilyMembers LY6G6C, LY6G6D, LY6G6E,LY6G5B and LY6G5C 244

15.2.4 Activating Natural Killer Receptor 24515.2.5 Butyrophilin-like Protein 2 24515.2.6 Advanced Glycosylation End-products

Receptor 24615.2.7 Leukocyte-specific Transcript 1 246

15.3 Genes that Encode Cytokines 24715.3.1 Tumour Necrosis Factor 24715.3.2 Lymphotoxin-a and -b 24815.3.3 Allograft Inflammatory Factor 1 248

15.4 Genes that Encode Proteins Involved in Response

to Stress 24915.4.1 70 kDa Heat Shock Proteins HSPA1A,

HSPA1B and HSPA1L 24915.4.2 Activating Transcription Factor 6b 25015.4.3 FK-506 Binding Protein-like 251

15.5 Genes that Encode Proteins Involved in

Protein Ubiquitination 25115.5.1 E3 Ubiquitin-protein Ligase

RNF5 25115.5.2 Scythe (BAT3) 252

15.6 Genes that Encode Proteins Involved in

Transcriptional Control 25315.6.1 Serine/Threonine Protein Kinase 19 25315.6.2 Negative Elongation Factor Polypeptide E 25315.6.3 Nuclear Factor of Kappa Light Chain Gene

Enhancer in B Cells Inhibitor-like 1(NFKBIL1) 254

15.6.4 Euchromatic Histone-lysineN-methyltransferase 2 254

15.7 Genes that Encoding Proteins Involved

in mRNA Processing 25515.8 Genes that Encode Proteins Involved in Cell

Signalling 25615.8.1 Neurogenic Locus Notch Homologue

Protein 4 25615.8.2 Casein Kinase II b Subunit 256

15.9 Genes that Encode Enzymes 25715.9.1 Sialidase 25715.9.2 G2 Subunit of Vacuolar H1-ATPase 257

15.10 Conclusions 258References 259

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Section 6 Hyaluronan-Binding Proteins in Inflammation

Chapter 16 Hyaluronan-Binding Proteins in Inflammation

Anthony J. Day, Charles D. Blundell,

David J. Mahoney, Marilyn S. Rugg and Caroline M. Milner

16.1 About the Authors 26916.2 Introduction and Historical Perspective 27116.3 Structural and Molecular Studies

on Hyaluronan-Binding Proteins 27116.3.1 TSG-6 and Type A HABD 27316.3.2 Type C HABD in the Link Proteins and

Lecticans 27616.3.3 Type B HABD of CD44 277

16.4 Investigating TSG-6 Biology 27916.4.1 TSG-6 in Ovulation 28016.4.2 TSG-6 in Arthritis and Inflammatory

Disease 28416.5 Summary and Final Thoughts 285Acknowledgements 286References 286

Subject Index 291

xiv Contents