title: Proteolytic processing of the astrovirus capsid.
abstract: To further characterize the nature of proteolytic processing of the astrovirus capsid, we infected Caco-2 cells with a high multiplicity of astrovirus without trypsin in the presence of 5 to 10% fetal calf serum. These infections were characterized by pulse-chase labeling with [35S]Smethionine, electron microscopy, gel electrophoresis of purified viral particles, and analysis of infectivity of such particles with and without added trypsin. Pulse-chase experiments showed that the astrovirus capsid protein was initially translated as an approximately 87-kDa protein. The 87-kDa capsid protein was rapidly converted intracellularly to a 79-kDa form which was found in smaller amounts in the cell supernatant. Purification by differential centrifugation yielded particles that appeared quite similar to trypsin-grown astrovirus particles by negatively stained electron microscopy. These particles were antigenically distinct from trypsin-treated virions as demonstrated by their various reactions with monoclonal antibodies in a solid-phase immunoassay. The purified trypsin-free particles were mainly composed of the 79-kDa capsid protein which was found to have an amino terminus at residue 71 of the entire open reading frame 2 (ORF2) product. The cleavage site was identified in a highly conserved region of the astrovirus ORF2 product. These trypsin-free particles were minimally infectious in cultured Caco-2 cells but became highly infectious (10(5)-fold increase) after trypsin but not chymotrypsin treatment. This trypsin-enhanced infectivity correlated with conversion of the 79-kDa capsid protein to three smaller peptides of approximately 34, 29, and 26 kDa.
authors.affiliation: Department of Pediatrics, Stanford University, Stanford, California 94305, USA. Dorsey.Bass@Forsythe.Stanford.edu
authors.forename: D M
authors.initials: DM
authors.lastname: Bass
authors.forename: S
authors.initials: S
authors.lastname: Qiu
mesh_terms: D000956:Antigens, Viral; D018938:Caco-2 Cells; D002213:Capsid / Q000378:metabolism*; D006801:Humans; D001257:Mamastrovirus / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D014357:Trypsin; D014771:Virion / Q000648:ultrastructure
chemical_list: D000956:Antigens, Viral; D011498:Protein Precursors; D014357:Trypsin