BIOPHYSICAL CHARACTERIZATION OF ZINC EJECTION FROM HIV NUCLEOCAPSID PROTEIN BY ANTI-HIV 2,2'-DITHIOBIS[BENZAMIDES] AND BENZISOTHIAZOLONES

HIV nucleocapsid protein (NCp7) has been suggested as a possible targe t for 2,2'-dithiobis[benzamide] and benzisothiazolone agents that inhi bit viral replication in infected cells (Rice et al. Science 1995, 270 , 1194-1197). The solution behavior of these compounds and the mechani stic events leading to removal of Zn from HIV nucleocapsid protein in vitro has been studied by electrospray ionization mass spectrometry, 5 00 MHz one- and two-dimensional nuclear magnetic resonance spectroscop y, and circular dichroism spectroscopy. We demonstrate that (1) Zn eje ction is accompanied by formation of covalent complexes formed between the 2,2'-dithiobis[benzamide] monomers and Cys residues of Zn-deplete d NCp7, (2) the rate of Zn ejection is faster for the C-terminal Zn fi nger and slower for the N-terminal finger, (3) Zn ejection results in a loss of structural integrity of the NCp7 protein, and (4) there is n o appreciable interaction between a nonreactive iso stere of the lead 2,2'-dithiobis[benzamide] and NCp7 in buffered aqueous solution. These findings are discussed in terms of the mechanism of action of Zn ejec tion by aromatic 2,2'-dithiobis[benzamides].