Design, synthesis and CD4 binding studies of a fluorescent analogue of a peptide that enhances HIV-1 infectivity.

ABSTRACT

We previously demonstrated that a 23-amino-acid peptide derived from the V3 loop of the surface glycoprotein of human immunodeficiency virus (HIV-1) strain MN was able to bind soluble CD4 and to enhance HIV-1 infection. Further studies suggested that the peptide/CD4 interaction induces an increase in both CD4 expression and CD4/gp120 binding affinity. To facilitate identification of the complementary binding site for the peptide on cellular CD4, we designed an analogue carrying a single fluorescein moiety. The synthesis of this modified analogue presented several problems because of the presence of several amino acids in the sequence carrying potentially reactive groups in their side-chains, and the necessity of introducing only one marker per molecule in a position that would not affect biological activity. The side-chain of Lys19 was selected because separate studies demonstrated that its substitution with an uncharged amino acid does not reduce the peptide's biological activity. We compared the merits of various synthetic protocols used to condense the fluorescent marker with the peptide. Biological assays indicated that the presence of the fluorescein moiety did not compromise peptide binding to CD4; furthermore, binding of the labeled analogue was not abolished by trypsin treatment, suggesting that the peptide may interact with both CD4 and additional trypsin-resistant binding sites on the cell surface. Finally, we verified the preservation of HIV infection enhancing ability in the labeled peptide.