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

¹ Department of Chemistry, University of Nebraska–Lincoln, Lincoln, Nebraska 68588, USA
² Department of Plant Pathology, University of Nebraska–Lincoln, Lincoln, Nebraska 68588, USA

Reprint requests to: Dr. David L. Smith, Department of Chemistry, University of Nebraska–Lincoln, Lincoln, Nebraska 68588-0304, USA; e-mail: dls{at}unlserve.unl.edu; fax: (402) 472-9402.

Amide hydrogen exchange and mass spectrometry have been used to study the pH-induced structural changes in the capsid of brome mosaic virus (BMV). Capsid protein was labeled in a structurally sensitive way by incubating intact 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 spectrometry. The largest deuterium increases induced by structural alteration occurred in the regions around the quasi-threefold axes, which are located at the center of the asymmetric unit. The increased levels of deuterium indicate loosening of structure in these regions. This observation confirms the previously proposed swelling model for BMV and cowpea chlorotic mottle virus (CCMV) and is consistent with the structure of swollen CCMV recently determined by cryo-electron microscopy and image reconstruction. Structural changes in the extended N- and C-terminal arms were also detected and compared with the results obtained with other swollen plant viruses. This study demonstrates that protein fragmentation/amide hydrogen exchange is a useful tool for probing structural changes in viral capsids.

Keywords: Brome mosaic virus; amide hydrogen exchange; mass spectrometry; viral capsid

Abbreviations: BMV, brome mosaic virus • CCMV, cowpea chlorotic mottle virus • TBSV, tomato bushy stunt virus • CP, capsid protein • EM, electron microscopy • ESI, electrospray ionization • GdHCl, guanidine hydrochloride • HX, hydrogen exchange • HPLC, high performance liquid chromatography • MS, mass spectrometry • m/z, mass-to-charge ratio • TFA, trifluoroacetic acid

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