Characterization of intermediates in the oxidation of zinc fingers in human immunodeficiency virus type 1 nucleocapsid protein P7
Hathout, Y.; Fabris, D.; Han, M.S.; Sowder, R.C.; Henderson, L.E.; Fenselau, C.
Drug Metabolism and Disposition the Biological Fate of Chemicals 24(12): 1395-1400
1996
ISSN/ISBN: 0090-9556 PMID: 8971148 Document Number: 464950
Oxidants targeted toward inactivation of the nucleocapsid zinc finger protein are under development as antiviral agents, especially for use against human immunodeficiency virus. In the present study, electrospray ionization-mass spectrometry is used to follow in situ the progress of the reactions of 2,2'-dithiodipyridine and disulfiram with recombinant nucleocapsid protein p7 (Ncp7) from human immunodeficiency virus-1 at pH 7.4. Both reagents react with the two zinc fingers in the protein, resulting in the ejection of two zinc ions and the formation of oxidized apo-Ncp7 with three intramolecular disulfide bonds. The ejection of zinc by 2,2'-dithiodipyridine occurs in two steps. Alkylation of unreacted cysteine residues with N-ethylmaleimide after a 2-min reaction with 2,2'-dithiodipyridine reveals that the carboxyl-terminal zinc finger is disrupted first. Cys-49, Cys-36, and, to a lesser extent, Cys-39 are all shown to be target residues for initial electrophilic attack. In the reaction of disulfiram with Ncp7, ejection of the two zinc ions also occurs in two steps; however, the fully oxidized apo-Ncp7 is formed more rapidly. Thus, after a 40-min reaction, 45% of native Ncp7 is oxidized by 2,2'-dithiodipyridine, whereas 75% is oxidized by disulfiram.