A SYNTHETIC CONFORMATIONAL EPITOPE FROM THE C4 DOMAIN OF HIV GP120 THAT BINDS CD4

The fourth conserved domain of the human immunodeficiency virus type 1 (HIV-1) envelope, the C4 region of glycoprotein 120 (gp120), is belie ved to be a major part of gp120 that is necessary for binding to CD4, Recently, we found that C4 in gp120 is probably an alpha-helix, becaus e antibodies made against helical constructs of C4 react with native a nd recombinant gp120 but antibodies against linear C4 constructs do no t. For the present study, we performed experiments to determine, first , if CD4 could bind to the helical C4 constructs and, second, if the b inding was comparable with CD4 binding to gp120. Immobilized helical c onstructs derived from the C4s from HTV-1 and HIV-2 bound biotinylated recombinant CD4 with K-d values of 8.59 nM and 14.59 nM, respectively . Recombinant soluble CD4 inhibited the binding of biotinylated CD4 to the C4 construct from HIV-1 with a K-d of 9.88 nM, and recombinant gp 120 blocked the binding of CD4 to the immobilized helical construct fr om C4 of HIV-1 with a K-d of 8.08 nn. The C4 peptide-(419-436) from HI V-1 (KIKQIINMWQEVGKAMYA-NH2) blocked CD4 binding to gp120 but only in a buffer con containing 0.03% Brij 35 where the peptide displayed 17 /- 1% alpha-helix; without the Brij 35, peptide-(419-436) displayed no helical content, The K-d for the peptide-(419-436) blocking CD4 bindi ng to gp120 in Brij 35-containing buffer was found to be 42 mu M. Thes e results indicate that C4 constructs from HTV-1 and HIV-2 do bind CD4 , but the constructs must display an alpha-helical conformation to do so. In addition, the results reported here will provide answers to key questions about structural requirements for HIV vaccines and therapeu tics that hinge on under standing the molecular nature of the gp120-CD 4 interaction as the first step in the HIV infection process.