Detecting structural changes in viral capsids by hydrogen exchange and mass spectrometry

Amide hydrogen exchange and mass spectrometry have been used to study the p H-induced structural changes in the capsid of brome mosaic virus (BMV). Cap sid protein was labeled in a structurally sensitive way by incubating intac t viral particles in D2O at pH 5.4 and 7.3. Deuterium levels in the intact coat protein and its proteolytic fragments were determined by mass spectrom etry. The largest deuterium increases induced by structural alteration occu rred in the regions around the quasi-threefold axes, which are located at t he center of the asymmetric unit. The increased levels of deuterium indicat e loosening of structure in these regions. This observation confirms the pr eviously proposed swelling model for BMV and cowpea chlorotic mottle virus (CCMV) and is consistent with the structure of swollen CCMV recently determ ined by cryo-electron microscopy and image reconstruction. Structural chang es in the extended N- and C-terminal arms were also detected and compared w ith the results obtained with other swollen plant viruses. This study demon strates that protein fragmentation/amide hydrogen exchange is a useful tool for probing structural changes in viral capsids.