Molecular Interaction Studies of HIV-1 Matrix Protein p17 and Heparin: IDENTIFICATION OF THE HEPARIN-BINDING MOTIF OF p17 AS A TARGET FOR THE DEVELOPMENT OF MULTITARGET ANTAGONISTS

Once released by HIV cells, p17 binds heparan sulfate proteoglycans
(HSPGs) and CXCR1 on leukocytes causing their
dysfunction. By exploiting an approach integrating computational
modeling, site-directed mutagenesis of p17, chemical
desulfation of heparin, and surface plasmon resonance, we characterized
the interaction of p17 with heparin, a HSPG structural
analog, and CXCR1. p17 binds to heparin with an affinity (Kd
190 nM) that is similar to those of other heparin-binding viral
proteins. Two stretches of basic amino acids (basic motifs) are
present in p17 N and C termini. Neutralization (Arg3Ala substitution)
of the N-terminal, but not of the C-terminal basic
motif, causes the loss of p17 heparin-binding capacity. The
N-terminal heparin-binding motif of p17 partially overlaps the
CXCR1-binding domain. Accordingly, its neutralization prevents
also p17 binding to the chemochine receptor. Competition
experiments demonstrated that free heparin and heparan
sulfate (HS), but not selectively 2-O-, 6-O-, and N-O desulfated
heparins, prevent p17 binding to substrate-immobilized heparin,
indicating that the sulfate groups of the glycosaminoglycan
mediate p17 interaction. Evaluation of the p17 antagonist activity
of a panel of biotechnological heparins derived by chemical
sulfation of the Escherichia coli K5 polysaccharide revealed that
the highlyN,O-sulfated derivative prevents the binding of p17 to
both heparin and CXCR1, thus inhibiting p17-driven chemotactic
migration of human monocytes with an efficiency that is
higher than those of heparin and HS. Here, we characterized at a
molecular level the interaction of p17 with its cellular receptors,
laying the basis for the development of heparin-mimicking p17
antagonists.