RSV entry inhibitors block F-protein mediated fusion with model membranes.
RSV fusion is mediated by F-protein, a major viral surface glycoprotein. CL-309623, a specific inhibitor of RSV, interacts tightly with F-protein, which results in a hydrophobic environment at the binding site. The binding is selective for F-protein and does not occur with G-protein, a surface glycoprotein that facilitates the binding of RSV to target cells, or with lipid membranes at concentrations in the sub-millimolar range. Using an assay based on the relief of self-quenching of octadecyl rhodamine (R18) incorporated in the RSV envelope, we show that the virus fuses efficiently with large unilamellar vesicles containing cholesterol, in the absence of specific receptor analogs. Fusion of cp-52, a mutant virus lacking the G and SH surface glycoproteins, with vesicles is inhibited by CL-309623 and RFI-641 due to specific interactions of the inhibitor(s) with the fusion protein. Both virus-vesicle and virus-cell fusion are inhibited with equal potency. The formation of the binary complex of CL-309623 with F-protein in its native state, resulting in the inhibition of fusion and entry of virus, is a prerequisite for the observed anti-RSV activity in cell cultures.
Razinkov
Vladimir
V
Department of Biological Chemistry, Wyeth Research, Pearl River, NY 10965, USA.
Huntley
Clayton
C
Ellestad
George
G
Krishnamurthy
Girija
G
D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002522:Chlorocebus aethiops; D005456:Fluorescent Dyes; D015707:HN Protein / Q000235:genetics / Q000378:metabolism; D050356:Lipid Metabolism; D009154:Mutation; D011485:Protein Binding / Q000187:drug effects; D018113:Respiratory Syncytial Virus, Human / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D012235:Rhodamines; D013449:Sulfonamides / Q000494:pharmacology*; D022161:Transport Vesicles / Q000378:metabolism; D014227:Triazines / Q000494:pharmacology*; D014709:Vero Cells; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects
D005456:Fluorescent Dyes; D015707:HN Protein; C434804:RFI 641; D012235:Rhodamines; D013449:Sulfonamides; D014227:Triazines; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; C116204:attachment protein G