FIG. 6.

Immunoblot analysis of E1 mutant proteins. Monolayers of induced BHK cells were lysed in lysis buffer containing 10 mM iodoacetamide to prevent the formation of disulfide bonds. The protein samples were denatured in the presence (+β-Me) or absence (−β-Me) of 2-mercaptoethanol, separated by SDS-PAGE, and transferred to nitrocellulose membranes for immunoblot analysis. RV antigens were detected by using human anti-RV serum (Human), anti-E1 monoclonal antibody exhibiting hemagglutination-inhibiting activity (Mab-E1), and anti-E2 monoclonal antibody (Mab-E2). The positions of the apparent molecular mass standards are indicated on the right (in kilodaltons). E1-E2 heterodimer is indicated by an asterisk. Wt, BHK-E2E1; Cys, BHK-E2E1(C82S); dt, BHK-E2E1(dt); Gly, BHK-E2E1(G93D); Pro, BHK-E2E1(P104G).