Structure and antigenicity of the divergent human astrovirus VA1 capsid spike
Fig 4
Processing and assembly of the mature HAstV-VA1 capsid protein.
(A) The complete amino acid sequence of the HAstV-VA1 capsid precursor protein (GenBank accession number YP_003090288.1) is shown. Amino acids highlighted in yellow were identified by mass spectrometry as peptides of VP33, and those highlighted in green correspond to peptides of the VP38. Amino acids that are not observed are highlighted in grey. The cleavage site that generates the amino-terminal end of VP38, previously determined by Edman degradation, is shown in bold. The blue boxes correspond to the N- and C-termini of the core domain, whereas the red boxes correspond to the N- and C-termini of the spike domain. The pink box corresponds to amino acid E296, the last amino acid detected in VP33. (B) Coomassie-stained SDS-PAGE analysis of CsCl-purified virus. Lane 1: molecular weight marker, in kD. Lane 2: two bands corresponding to the HAstV-VA1 capsid proteins VP33 and VP38. Bands were excised and utilized for proteolytic digestion, liquid chromatography, and tandem mass spectrometry to identify peptides. (C) Model of the mature HAstV-VA1 capsid protein. An AlphaFold2-predicted core domain structure, a linker region, and the spike domain crystal structure are shown, colored as in panel A. The second protomer of the spike domain is colored grey. The N-terminal amino acids 1–71 that are present in VP33 are not shown. Domain termini are labeled (R72 and P389 (core domain), and P408 and P680 (spike domain)). The location of the cleavage site that results in the N-terminus of VP38 (N347/T348) is indicated. The location of E296, the last observed amino acid of VP33, which is in a structurally similar site as a known trypsin cleavage site in classical HAstVs, is indicated. (D) Model of the mature classical HAstV-8 capsid protein. The crystal structure of the HAstV-8 core domain (PDB: 5IBV), a linker region, and the crystal structure of the HAstV-8 spike domain (PDB: 3QSQ) are shown. The second protomer of the spike domain is colored grey. The N-terminal amino acids 1–76 that are present in VP34 are not shown. Domain termini are labeled (R77 and L412 (core domain), and E424 and P645 (spike domain)). The location of the trypsin cleavage site that results in the N-terminus of VP27 (R393/Q394) is indicated. The location of R313, a trypsin cleavage site, is indicated.