Covalent interactions are not required to permit or stabilize the non-covalent association of hepatitis C virus glycoproteins E1 and E2

Hepatitis C virus (HCV) encodes two glycoproteins, E1 and E2, which are tho ught to locate to the envelope of virus particles. These proteins form two complexes in tissue culture systems, a high molecular mass aggregate that c ontains intermolecular covalent bonds and a native complex in which E1 and E2 associate by non-covalent interactions, The contribution of either compl ex to the structures of the proteins on virus particles is not known. Using dithiothreitol to reduce inter- and intramolecular disulphide bonds in sit u, we have studied the nature of the interactions within the aggregate and the role of covalent bonds in the early stages of E1-E2 association, Result s with two HCV type la strains, Glasgow and H77, showed that the aggregate contains not only covalent interactions but also non-covalent associations between E1 and E2, These non-covalent associations are complex since deleti on mutant analysis failed to identify any single region which was required for non-covalent interaction, Complex formation by de novo synthesized prot eins was not arrested under reducing conditions which prevented the product ion of inter- and intramolecular disulphide bonds, Moreover, at conformatio n-specific antibody continued to recognize the E2 protein in reduced comple xes, indicating that covalent bonds do not stabilize certain structures of E2 that can interact with E1. These data suggest that disulphide bonds are not required either to allow association between the proteins or to stabili ze E1-E2 complexes.