The catalytic activity of FBL is required for HeV infection but not RSV infection.
(A) Crystal structure of human FBL (RCSP Protein Databank 2IPX) in complex with 5‘-deoxy-5'-methylthioadenosine (MTA) (red), an analog of the natural cosubstrate factor S-adenosylmethionine (SAM). (http://www.rcsb.org/pdb/explore/explore.do?structureId=2IPX). The two conserved residues critical for co-substrate binding are shown. Red lines depict polar contacts between MTA and the huFBL molecule. E191 (orange) and D236 (green) were each substituted with alanine. (B) Protein expression levels of FBL variants assessed by Western blotting of HeLa cell lysates, using an anti-FLAG antibody. HeV infection (C) and RSV infection (D) in HeLa cells depleted of endogenous FBL +/- a vector (pCMV2, 500 ng, 24 h) expressing FBL resistant to siRNA carrying the E191A mutant, followed by HeV infection (MOI 0.1, 24 h). *p<0.05 compared to vector.