Crystal structure of raptor adenovirus 1 fibre head and role of the beta-hairpin in siadenovirus fibre head domains

Abstract

Background: Most adenoviruses recognize their host cells via an interaction of their fibre head domains with a primary receptor. The structural framework of adenovirus fibre heads is conserved between the different adenovirus genera for which crystal structures have been determined (Mastadenovirus, Aviadenovirus, Atadenovirus and Siadenovirus), but genus-specific differences have also been observed. The only known siadenovirus fibre head structure, that of turkey adenovirus 3 (TAdV-3), revealed a twisted beta-sandwich resembling the reovirus fibre head architecture more than that of other adenovirus fibre heads, plus a unique beta-hairpin embracing a neighbouring monomer. The TAdV-3 fibre head was shown to bind sialyllactose.

Methods: Raptor adenovirus 1 (RAdV-1) fibre head was expressed, crystallized and its structure was solved and refined at 1.5 Å resolution. The structure could be solved by molecular replacement using the TAdV-3 fibre head structure as a search model, despite them sharing a sequence identity of only 19 %. Versions of both the RAdV-1 and TAdV-3 fibre heads with their beta-hairpin arm deleted were prepared and their stabilities were compared with the non-mutated proteins by a thermal unfolding assay.

Results: The structure of the RAdV-1 fibre head contains the same twisted ABCJ-GHID beta-sandwich and beta-hairpin arm as the TAdV-3 fibre head. However, while the predicted electro-potential surface charge of the TAdV-3 fibre head is mainly positive, the RAdV-1 fibre head shows positively and negatively charged patches and does not appear to bind sialyllactose. Deletion of the beta-hairpin arm does not affect the structure of the raptor adenovirus 1 fibre head and only affects the stability of the RAdV-1 and TAdV-3 fibre heads slightly.

Conclusions: The high-resolution structure of RAdV-1 fibre head is the second known structure of a siadenovirus fibre head domain. The structure shows that the siadenovirus fibre head structure is conserved, but differences in the predicted surface charge suggest that RAdV-1 uses a different natural receptor for cell attachment than TAdV-3. Deletion of the beta-hairpin arm shows little impact on the structure and stability of the siadenovirus fibre heads.

Keywords: Atomic structure; Beta-hairpin; Deletion mutagenesis; Protein stability; Siadenovirus; X-ray crystallography.

Fig. 1

Structure of the RAdV-1 fibre head domain and comparison with the TAdV-3 fibre head. a The RAdV-1 fibre head monomer. Beta-strands are labelled (except the J'-strand, which is located behind the A-, B- and C-strands). The beta-hairpin is marked with an asterisk and the short helical region in the CD-loops with an alpha sign. b The RAdV-1 fibre head trimer. The three monomers are shown in green, magenta and cyan. The beta-hairpin and helical region are marked as in panel (a). c The RAdV-1 fibre head monomer (green; in the same orientation as in panel a) superposed onto the TAdV-3 fibre head monomer (dark blue). d The RAdV-1 fibre head trimer, coloured as in panel b, superposed onto the TAdV-3 fibre head trimer (dark blue). e Structure-based sequence alignment of the RAdV-1 and TAdV-3 fibre heads. Identical residues are indicated with lines, similar residues with dots. Beta-strands and the short helical region are highlighted. The residues that in the TAdV-3 fibre head structure interact with sialyllactose are underlined

Fig. 2

Electrostatic surface potential of the RAdV-1 and TAdV-3 fibre head domains. a The RAdV-1 fibre head viewed from the top. b The RAdV-1 fibre head monomer viewed from the side. The end of the beta-hairpin is marked with an asterisk. c The TAdV-3 fibre head (PDB entry 4D62) viewed from the top. d The TAdV-3 fibre head viewed from the side. The end of the beta-hairpin is marked with an asterisk and the 2,3-sialyllactose ligand is shown in stick representation (the ligand is hidden when viewed from the top)

Fig. 3

Thermal stability of siadenovirus fibre heads. a Thermofluor protein denaturation assay of the RAdV-1 fibre head (with beta-hairpin, continuous line) and RAdV-1 fibre head from which the beta-hairpin arm residues 359-373 have been deleted (dotted line). b Thermofluor protein denaturation assay of the TAdV-3 fibre head (with beta-hairpin, continuous line) and TAdV-3 fibre head from which the beta-hairpin arm residues 349-364 have been deleted (dotted line)

Fig. 4

Superposition of the native and mutant RAdV-1 fibre head domain structures. a The RAdV-1 fibre head monomer in native (with the beta-hairpin arm; green) and mutant form (without the beta-hairpin arm; yellow), viewed from the side. b The RAdV-1 fibre head trimer in native (with the beta-hairpin arm; monomer in green, magenta and cyan) and mutant form (without the beta-hairpin arm; in yellow), viewed from the top. The end of the beta-hairpin of monomer A is marked with an asterisk in both panels