UC Berkeley

Viral fusion proteins are essential for most enveloped viruses to infect cells, making them key targets for vaccines and therapeutics. Despite a wealth of structural data, their conformational dynamics remain insufficiently understood, presenting challenges to optimizing vaccine and therapeutic design. Viral fusion proteins undergo dramatic conformational changes during fusion, making it both an interesting protein folding question and a therapeutic opportunity. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) is uniquely suited to investigate the conformational landscapes of viral fusion proteins, providing insights into solvent accessibility and secondary structure stability under diverse conditions.This dissertation employs HDX-MS to study the conformational dynamics of the SARS-CoV-2 spike protein and Rabies virus glycoprotein (RABV-G). Chapter 1 reviews viral fusion proteins and their study. Chapter 2 discusses our work that revealed a novel, open-interface trimer conformation of the SARS-CoV-2 spike protein, exposing previously inaccessible surface areas to solvent and antibodies. Temperature, receptor binding, and sequence variations influence the thermodynamics and kinetics of this transition. Chapter 3 extends these findings by demonstrating that this conformation is sampled on enveloped virus-like particles (eVLPs) and allows us to see how other engineering efforts and natural evolution have shaped the conformational landscape of spike. This chapter also reveals that cleavage at the multibasic furin site allosterically increases the flexibility of the S2’ protease site — providing a molecular basis for enhanced infectivity in viruses with this cleavage site. Chapter 4 explores ongoing studies of RABV-G, comparing its conformational dynamics as a soluble ectodomain and on eVLPs, highlighting the membrane's role in shaping its conformational landscape. These findings deepen our understanding of viral fusion protein dynamics, offering molecular insights to inform vaccine and therapeutic design.