Influenza A Virus Hemagglutinin: An Examination of Divergent Subtypes with Respect to Stability and Receptor Binding Properties Público
Byrd-Leotis, Lauren Ashley (2016)
Published
Abstract
The hemagglutinin (HA) of influenza A viruses mediates the processes associated with viral entry: binding to cell surface receptors and facilitating fusion of the virion membrane with the host endosomal membrane. The HA structure incorporates two functional domains related to these activities, the receptor binding site in the globular head and the fusion peptide and pocket in the stem region. These sites have specific characteristics associated with subtype and host, and the differences often define species barriers. In this work, we have characterized the effects of mutations in the stem region of HA for strains representing each structural group, as well as avian and human hosts. We found that for the conserved residue 58, a lysine to isoleucine change increased the acid stability of most HAs, lowering the pH of fusion. A mutation at conserved residue 112, from aspartic acid to glycine, decreased the acid stability, raising the pH of fusion for all HA subtypes. The effects of both mutations were consistent across structural and host classifications, correlating with the conserved nature of both residues. Future studies are planned to examine the effects of mutations at group specific positions. We began our study of receptor binding by characterizing the broad receptor specificity of a range of influenza viruses again differing in subtype and host. The receptor specificity is linked to residues within the receptor binding pocket of HA, though their positions are altered between group 1 and group 2 subtypes. Avian viruses preferentially recognize α2,3-Sialic acid terminating glycans and mammalian viruses recognize α2,6-Sialic acid. Our results using the Consortium for Functional Glycomics glycan microarray corroborate these statements, though they reveal a general over-simplification in that other structural determinants beyond the terminating sialic acid moiety are important for Influenza A virus binding. To identify the receptors used in natural infection, we developed the Pig Lung Shotgun Microarray incorporating N-glycans isolated from swine lung tissue. Eight endogenous receptors were identified and characterized. Continuation of this work involves expanding our repertoire of natural arrays to include human lung glycans and examining the role of neuraminidase in entry as the substrate specificity overlaps with that of HA.
Table of Contents
ABSTRACT
ACKNOWLEDGMENTS
TABLE OF CONTENTS
LIST OF FIGURES AND TABLES
INTRODUCTION 1
The Virus Life Cycle and Management of Infection 3
Receptor Binding and Entry 3
Transcription 7
Translation 8
Assembly and Budding 9
Vaccines and Antivirals 10
Subtypes, Reassortment, and Pandemic Events 12
The Role of the Host: Carbohydrate Interaction, Interspecies Transmission, And Pathogenicity 16
Carbohydrates Recognized by Influenza A Virus 16
Getting to The Surface- Mucins and Virion Morphology 18 Presentation of Receptors in Avian, Swine, and Human Hosts 21
Interspecies Transmission 26
Pathogenicity 29
Functional Balance of HA and NA 30
Dissertation Overview 32
INFLUENZA HEMAGGLUTININ (HA) STEM REGION MUTATIONS THAT STABILIZE OR DESTABILIZE THE STRUCTURES OF MULTIPLE HA SUBTYPES 34
Abstract 35
Importance 36
Introduction 36
Materials and Methods 40
Results 45
Discussion 55
Acknowledgments 61
SHOTGUN GLYCOMICS OF PIG LUNG IDENTIFIES NATURAL ENDOGENOUS RECEPTORS FOR INFLUENZA VIRUSES 62
Abstract 63
Significance Statement 64
Introduction 65
Results 70
Discussion 85
Experimental Methods 92
Acknowledgments 97 Supplementary Material 98
CONCLUSIONS AND FUTURE STUDIES 116
APPENDIX: ANALYSIS OF FUSION PEPTIDE POCKET RESIDUES OF INFLUENZA A VIRUS HEMAGGLUTININ AND THEIR EFFECT ON THE pH STABILITY OF THE PROTEIN 125
Abstract 126
Introduction 127
Materials and Methods 129
Results and Discussion 134
Acknowledgments 142
REFERENCES 146
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Influenza A Virus Hemagglutinin: An Examination of Divergent Subtypes with Respect to Stability and Receptor Binding Properties () | 2018-08-28 12:15:29 -0400 |
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