HEPATITIS-C VIRUS ENVELOPE PROTEINS BIND LACTOFERRIN

Hepatitis C virus (HCV) has two envelope proteins, E1 and E2, which fo rm a heterooligomer, During dissection of interacting regions of HCV E 1 and E2, we found the presence of an interfering compound or compound s in skim milk, Here we report that human as well as bovine lactoferri n, a multifunctional immunomodulator, binds two HCV envelope proteins. As determined by far-Western blotting, the bacterially expressed E1 a nd E2 could bind lactoferrin in human milk directly separated or immun opurified and separated by sodium dodecyl sulfate-polyacrylamide gel e lectrophoresis, The bindings of lactoferrin and HCV envelope proteins in vitro were confirmed by another method, the pull-down assay, with i mmunoprecipitated lactoferrin-bound protein A resin, By the same assay , mammal-expressed recombinant E1 and E2 were also demonstrated to bin d human lactoferrin efficiently in vitro, Direct interaction between E 2 and lactoferrin was proved in vivo, since anti-human lactoferrin ant ibody efficiently coimmunoprecipitated with secreted and intracellular forms of the E2 protein, but not glutathione S-transferase (GST), fro m lysates of HepG2 cells transiently cotransfected with the expression plasmids of human lactoferrin and gE2t-GST (the N-terminal two-thirds of E2 fused to GST) or GST, The N-terminal loop of lactoferrin, the r egion important for the antibacterial activity, has only a little role in the binding ability to HCV E2 but affected the secretion or stabil ity of lactoferrin, Taken together, these results indicate the specifi c interaction between lactoferrin and HCV envelope proteins in vivo an d in vitro.