STRUCTURAL AND FUNCTIONAL-ANALYSIS OF NEGATIVELY CHARGED MILK-PROTEINS WITH ANTI-HIV ACTIVITY

Several polyanionic reagents such as dextran sulfates, heparin sulfate s, and negatively charged proteins have been reported to exhibit anti- HIV activity in vitro, Particularly potent inhibition has been reporte d for the milk protein beta-lactoglobulin (beta LG) on modification by 3-hydroxyphthalic anhydride (3HP), The introduction of multiple negat ively charged carboxyl groups along the polypeptide backbone obviously leads to repulsion within the protein molecule and this is likely to affect the specific tertiary, and perhaps also secondary, structure of the protein, We used several biophysical techniques to probe the stru ctural changes that occur on 3HP modification of beta LG, The results suggest that the protein becomes largely unstructured on chemical modi fication, Although a profound anti-HIV activity was measured for 3HP-b eta LG, similar antiviral effects were observed with two other 3HP-mod ified milk proteins, alpha-lactalbumin and alpha(S2)-casein, but not w ith the unmodified proteins, Most potent inhibition of HIV-1 replicati on was obtained with 3HP-modified alpha-lactalbumin, which also demons trated the least cytotoxicity, These combined results indicate that HI V inhibition is a general property of negatively charged polypeptides and do not support a model in which the negatively charged 3HP-beta LG protein interacts in a structure-specific manner with the CD4 cell su rface receptor for HIV-1 entry.