Structure-function analysis of hepatitis C virus envelope-CD81 binding

Hepatitis C virus (HCV) is a major human pathogen causing chronic liver dis ease. We have recently found that the large extracellular loop (LEL) of hum an CD81 binds HCV. This finding prompted us to assess the structure-functio n features of HCV-CD81 interaction by using recombinant E2 protein and a re combinant soluble form of CD81 LEL. We have found that HCV-E2 binds CD81 LE L with a K-d of 1.8 nM; CD81 can mediate attachment of E2 on hepatocytes; e ngagement of CD81 mediates internalization of only 30% of CD81 molecules ev en after 12 h; and the four cysteines of CD81 LEL form two disulfide bridge s, the integrity of which is necessary for CD81-HCV interaction. Altogether our data suggest that neutralizing antibodies aimed at interfering with HC V binding to human cells should have an affinity higher than 10(-9) M, that HCV binding to hepatcytes may not entirely depend on CD81, that CD81 is an attachment receptor with poor capacity to mediate virus entry, and that re ducing environments do not favor CD81-HCV interaction. These studies provid e a better understanding of the CD81-HCV interaction and should thus help t o elucidate the viral life cycle and to develop new strategies aimed at int erfering with HCV binding to human cells.