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Hydrogen exchange mass spectrometry studies of structure and dynamics of viral capsids
Most viruses consist of nucleic acid encapsulated in a protein shell or capsid. Viral capsids are highly organized and dynamic protein assemblies that play a major role in viral infectivity. Hydrogen exchange mass spectrometry (HX MS) was used to study the structure and dynamics of viral capsids. Structural changes in the capsid of brome mosaic virus induced by pH changes were characterized by comparing deuterium levels in the capsid labeled at pH 5 and 7. A general destabilization was found at the higher pH, indicated by the increased levels of deuterium in the capsid protein. Peptic fragmentation of the labeled capsid protein showed substantially reduced stability in the region surrounding the quasi three-fold axis at the center of the icosahedral trimer. In contrast, the C-terminus and its surrounding pocket in an adjacent subunit retained a high level of stability. ^ To study a more complex animal virus, the HX MS procedure was optimized in terms of enzymatic digestion and sensitivity using fully deuterated cytochrome c. An immobilized pepsin column was used to improve protein digestion efficiency at 0°C. Capillary HPLC was used to scale down the chromatography and thus enhance the HX MS sensitivity by 100-fold while maintaining high amide deuterium recoveries. The improved HX MS method was used to study the human rhinovirus 14 capsid, which is composed of 4 different coat proteins VP1–4. Continuous labeling of the intact virus revealed a higher deuterium level in VP4 than VP1–3, consistent with an extended conformation of VP4 in contrast with a compact β-barrel fold in VP1–3. Deuterium levels in peptic fragments of VP1–4 were correlated with the secondary structures, B-factors, solvent accessibility and hydrogen bonding in these proteins. The dynamics of the protein-protein interface of a protomer and a pentamer was investigated by looking at exchange rates in these regions. These results facilitate future studies of the affects of antiviral agents on the structure and dynamics of HRV14 capsid. ^
Chemistry, Analytical|Chemistry, Biochemistry
Wang, Lintao, "Hydrogen exchange mass spectrometry studies of structure and dynamics of viral capsids" (2002). ETD collection for University of Nebraska - Lincoln. AAI3064573.