Investigating Protein and Peptide Mediated Membrane Bending Mechanisms Pubblico

Abstract

The ability to control and shape lipid bilayer is key to the survival of organisms, and they have evolved mechanisms to use membrane proteins as a driving force to bend membranes. In cells, membrane proteins such as the SNARE complex generates membrane curvature in synaptic vesicles such that neurotransmitter signals can be relayed. In influenza A, the pH-driven structural rearrangement of hemagglutinin is vital to fusing the viral and the host membranes and releasing ribonucleoprotein complexes into the host cell cytoplasm. The ability to remodel lipid bilayer is also essential to immune defense across living organisms. Particularly, host-defense peptides kill pathogens or viruses by folding into secondary structures on lipid membranes and ultimately disrupting pathogenic membranes or viral envelopes. This dissertation aims to study how protein and peptide conformational changes remodel lipid membranes. We explore this question from three angles: 1) the protein crowding mechanism, 2) influenza A membrane fusion, and 3) disruption of the membrane by a host-defense peptide. Together, these studies provide a better understanding of membrane bending mechanisms and the interactions between lipid and stimuli-responsive peptides.

Table of Contents

Chapter 1: Introduction to Protein and Peptide Mediated Membrane Bending

1.1 The Importance of Generating Curvature in Lipid Membranes………………………………...1

1.2 Mechanisms to Generate Curvature by Membrane Proteins……………………………………2

1.3 Example of Protein-Mediated Membrane Bending: Influenza A Viral Invasion………………...4

1.4 Example of Peptide-Mediated Membrane Bending: Host-Defense Peptides……………………8

1.5 Aims and Scope of Discussion…………………………………………………………………9

1.6 References…………………………………………………………………………………….11

Chapter 2: Peripheral Protein Unfolding Drives Membrane Bending

2.1 Abstract………………………………..……………………………………………………...16

2.2 Introduction………………………………..…………………………………………………17

2.3 Experimental Procedures……………………………..……………………………………….18

2.4 Results and Discussion……………………………..…………………………………………24

2.5 Conclusion……………………………..……………………………………………………...35

2.6 References……………………………..……………………………………………………....35

2.7 Supplemental Information……………………………..……………………………………...40

Chapter 3: Stability of HA2 Pre-fusion Structure Revealed by Sub-Millisecond Conformational Changes in the HA2 Domain of H3N2 Hemagglutinin

3.1 Abstract………………………………..……………………………………………………....60

3.2 Introduction………………………………..………………………………………………….60

3.3 Experimental Procedures………….………………..…..……………………………………...63

3.4 Results and Discussion……………………………..………………………………………….67

3.5 Conclusion……………………………..……………………………………………………...76

3.6 References……………………………..……………………………………………………....77

3.7 Supplemental Information……………………………..……………………………………....80

Chapter 4: pH-Dependent Liposome Crosslinking by Hemagglutinin HA2 B-Loop

4.1 Abstract………………………………..……………………………………………………....82

4.2 Introduction………………………………..………………………………………………….82

4.3 Experimental Procedures……………………………....………………………………..…..…84

4.4 Results and Discussion……………….…………..……...…………………………………….88

4.5 Conclusion……………………………..…………………………………………………......101

4.6 References……………………………..……………………………………………………..102

4.7 Supplemental Information……………………………..……………………………………..107

Chapter 5: Characterization of a Host-Defense Peptide, Urumin, and Its Interactions with Lipid membranes and the Influenza H3N2 X-31 Hemagglutinin

5.1 Abstract………………………………..……………………………………………………..114

5.2 Introduction………………………………..………………………………………………...115

5.3 Experimental Procedures………………………………………………..…………….……...117

5.4 Results and Discussion……………………………………………………………….……....120

5.5 Conclusion……………………………………………………………….……………….......131

5.6 References……………………………………………………………….……………….......132

5.7 Supplemental Information……………………………………………….……………….......136

Chapter 6: Conclusion

6.1 Summary………………………………..……………………………………………………137

6.2 Other Contributions………….……..………………………………………………………..138

6.3 References……………………………………………………………….……………….......138

About this Dissertation

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School	Laney Graduate School
Department	Chemistry
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Chemistry, Physical Chemistry, Biochemistry Biophysics, General
Parola chiave	membrane curvature protein folding kinetics influenza spectroscopy membrane protein-lipid interactions
Committee Chair / Thesis Advisor	R. Brian Dyer, Emory University
Committee Members	Khalid Salaita, Emory University James Kindt, Emory University

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