Vaccinia virus (VACV) belonging to the poxvirus family enters the host cell via two different entry pathways; either endocytosis or virus/host cell membrane fusion. With respect to the virus/host cell membrane fusion, there are eleven viral membrane proteins forming a complicated entry-fusion complex (EFC), including A28, A21, A16, F9, G9, G3, H2, J5, L5, L1 and O3, to conduct the fusion function. These EFC components are highly conserved in all poxviruses and each of them is essential and necessary for the fusion activity. So far, with the exceptions of L1 and F9 whose crystal structures were reported, the structural information about other EFC components remains largely unclear. We aim to conduct a structural and functional investigation of VACV virus-entry membrane protein A28. In this work, we expressed and purified a truncated form of A28 (14 kDa; residues 38-146, abbreviated as tA28 hereinafter), with deletion of its transmembrane domain (residues 1-22) and a hydrophobic segment (residues 23-37). And the assignments of its backbone and side chain 1H, 13C and 15N chemical shifts of tA28 are reported. The secondary structure propensity from TALOS+ indicates that tA28 does contain three α-helices, six β-strands and connecting loops. Aside from this, we demonstrated that tA28 does interact with fusion suppressor viral protein A26 (residues 351-500) by the 1H-15N HSQC spectrum. We interpret that A28 binding to A26 deactivates EFC fusion activity. The current study provides a valuable framework towards further structural analyses of this protein and for better understanding virus/host cell membrane fusion mechanism in association with virus entry.
Keywords: Chemical shift propensity; Entry-fusion complex; NMR resonance assignments; Vaccinia virus; Viral membrane protein.