SYNTHESIS AND CONFORMATIONAL STUDIES OF A CYCLIC ANALOG OF THE PROXIMAL ZINC-FINGER OF HIV-1 NCP7 FOR ANTIBODY GENERATION

The nucleocapsid protein NCp7 of human immunodeficiency virus type 1 (HIV-1) contains two zinc finger domains (CCHC boxes), which have been shown to be very important in the viral replication cycle. However, these domains are not involved directly in either in vitro RNA dimerization or tRNA(Lys,3) annealing. For a more detailed understanding of the role of the zinc fingers in the different functions of NCp7, antibodies directed against these domains would be very useful. For this purpose, a cyclic peptide analog of the proximal zinc finger (13-30)NCp7 has been synthesized in solid phase using a strategy of combined Fmoc and Boc chemistry. On the basis of the 3D structural data of NCp7. the Asn(17) and Ala(30) have been changed to Glu(17) and Lys(30) and a cyclization carried out between their side chains. The structures of the cyclic and native peptides complexed with Co2+ and Zn2+ were studied by visible and 2D H-1 NMR spectroscopy, respectively. The nuclear Overhauser effects obtained were applied as constraints to determine the solution structures using DIANA software followed by AMBER energy refinement. The results show that the cyclic peptide retains the highly folded structure of the native peptide and exhibits an enhanced affinity for metallic ions. These are favorable parameters for the generation of antibodies against the zinc fingers in NCp7.