Fusion Peptide of Influenza Hemagglutinin Requires a Fixed Angle Boomerang Structure for Activity

Judith M. White,Lukas K. Tamm,Heather Park,Alex L. Lai

doi:10.1074/jbc.m512280200

Abstract

The fusion peptide of influenza hemagglutinin is crucial for cell entry of this virus. Previous studies showed that this peptide adopts a boomerang-shaped structure in lipid model membranes at the pH of membrane fusion. To examine the role of the boomerang in fusion, we changed several residues proposed to stabilize the kink in this structure and measured fusion. Among these, mutants E11A and W14A expressed hemagglutinins with hemifusion and no fusion activities, and F9A and N12A had no effect on fusion, respectively. Binding enthalpies and free energies of mutant peptides to model membranes and their ability to perturb lipid bilayer structures correlated well with the fusion activities of the parent full-length molecules. The structure of W14A determined by NMR and site-directed spin labeling features a flexible kink that points out of the membrane, in sharp contrast to the more ordered boomerang of the wild-type, which points into the membrane. A specific fixed angle boomerang structure is thus required to support membrane fusion.

Highlights

To build a single fusion site, it is likely that multiples of three fusion peptides insert into the target membrane to initiate fusion between the viral and the target membrane
An assessment of the structures of the fusion peptides and transmembrane domains before, during, and after fusion is of high interest in our quest to understand the mechanism of influenza HA-mediated membrane fusion at the molecular level
Fusion Activity of HA Mutants—The angle of the boomerang of the fusion peptide structure in membranes is stabilized by hydrogen bonds and hydrophobic interactions between bulky apolar residues distributed around Asn-12, which forms the apex of the structure

Summary

Introduction

To build a single fusion site, it is likely that multiples of three fusion peptides insert into the target membrane to initiate fusion between the viral and the target membrane. The C-terminal half is more amphipathic with a number of polar and charged residues on its upper face and bulky hydrophobic residues including Trp-14 on its lower membrane-dipped face. This structure together with the close proximity of the N and C termini in the low pH crystal structure of the ectodomain and the lower energy of the pH 5 compared with the pH 7 conformation have led to the formulation of the spring-loaded boomerang model of influenza HA-mediated membrane fusion [6]. Our structure-energetics-function correlations support the notion that a specific fixed angle boomerang is required for tight interactions with lipids of the target membrane and for membrane fusion

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Conclusion