Research Projects

Membranes and Membrane Proteins

Membrane proteins represent approximately a third of expressed proteins in cells, yet they are under-represented in structural databases. We use solution and solid-state NMR to characterize the structures and dynamics of membrane proteins, and the curvature and fusion of membranes.

Membrane Fusion

Membrane fusion is ubiquitous in the biology of membranes and integral to neurotransmission, vesicle trafficking, cellular communication and viral infection. Membrane fusion is a non-spontaneous process with high-energy and highly curved intermediates that are catalyzed by fusion proteins. Our laboratory investigates the mechanism of membrane fusion in viral infection and neurotransmission.

Membrane Curvature

Curvature is inherent to the morphology of membranes, and it underpins the mechanism of membrane fusion. Certain organelles, such as the endoplasmic reticulum and the cristae of mitochondria, have highly curved membranes. Cells regulate membrane curvature with proteins and with lipid composition, utilizing lipids such as cardiolipins.

Previous studies focus on changes in the bulk properties of membranes, losing important information on the curvature effects of individual membrane components or the cooperativity of membrane curvature. We use NMR to measure membrane curvature and answer new questions on localized and microscopic membrane curvature, curvature cooperativity and changes in membrane structure with native lipids from eukaryotic tissues.

Viral Infection Proteins and Mechanisms

Influenza, HIV and Ebola viruses have an outer membrane envelope that must fuse with the host cell membrane on cellular entry. We use NMR to characterize the structures and mechanism of viral fusion proteins and the role of membrane curvature.

Viral Membrane Fusion

The influenza virus is associated with ca. 250,000-500,000 deaths yearly. It is a well-studied system, yet many questions remain in its molecular mechanism of transmission and infection. The hemagglutinin glycoprotein of the influenza A virus is a simple, prototypic membrane fusion system that fuses viral and host-cell membranes on cellular entry. The fusion peptide domain (HAfp) is vital to the function of the hemagglutinin, as it is highly conserved and interacts directly with the host membrane. HAfp peptides alone can fuse vesicles, and it is a useful model for membrane fusion. Previous approaches investigated truncated HAfp peptides that were partially unfolded, highly dynamic and significantly less fusiogenic. We solved the first structure of the full-length HAfp, revealing a new helical-hairpin amphipathic fold.

Our recent work ties this structure to the membrane fusion mechanism. Wildtype HAfp adopts a wedge structure that promotes negative curvature in membranes. We are additionally investigating the structures and mechanism of viral fusion proteins from viruses with similar fusion proteins, including the HIV, Ebola, MERS and SARS viruses, as well as the SNARE proteins involved in neurotransmission.

High-Resolution NMR Methods Development

Solution and solid-state NMR report detailed information on the distribution of electrons, the distances between atoms and the dynamics of molecules at the atomic level. We investigate new methods to increase the resolution of NMR structures, molecular dynamics and the distribution of conformation ensembles.

Hybrid NMR

Developments in NMR have focused on increasing the resolution of average structures. However, methods to characterize conformational ensembles and dynamics are limited, as they rely on changes in isotropic chemical shifts at hinge sites or the collection of data using numerous samples.

Hybrid NMR (hdNMR) is a new area of research we have developed for the study of biomolecular ensembles and dynamics. It combines the strengths and resolution of solution NMR with the detailed information of the anisotropic Hamiltonian from solid-state NMR. Its strength lies in the accurate characterization of chemical shift anisotropy (CSA) tensors in the aqueous state and the study of conformational distributions.