The role of Thr139 in the human immunodeficiency virus type 1 reverse transcriptase sensitivity to (+)-calanolide A

被引:14
作者
Auwerx, J
Rodríguez-Barrios, F
Ceccherini-Silberstein, F
San-Félix, A
Velázquez, S
De Clercq, E
Camarasa, MJ
Perno, CF
Gago, F
Balzarini, J
机构
[1] Katholieke Univ Leuven, Rega Inst Med Res, B-3000 Louvain, Belgium
[2] Univ Alcala de Henares, Dept Pharmacol, Alcala De Henares, Spain
[3] Univ Roma Tor Vergata, Dept Expt Med, Rome, Italy
[4] CSIC, Inst Quim Med, E-28006 Madrid, Spain
关键词
D O I
10.1124/mol.105.012351
中图分类号
R9 [药学];
学科分类号
1007 ;
摘要
The coumarins represent a unique class of non-nucleoside reverse transcriptase inhibitors (NNRTIs) that were isolated from tropical plants. (+)-Calanolide A, the most potent compound of this class, selects for the T139I resistance mutation in HIV-1 reverse transcriptase (RT). Seven RTs mutated at amino acid position 139 (Ala, Lys, Tyr, Asp, Ile, Ser, and Gln) were constructed by site-directed mutagenesis. The mutant T139Q enzyme retained full catalytic activity compared with wild-type RT, whereas the mutant T139I, T139S, and T139A RTs retained only 85 to 50% of the activity. Mutant T139K, T139D, and T139Y RTs had seriously impaired catalytic activities. The mutations in the T139I and T139D RTs were shown to destabilize the RT heterodimer. (+)-Calanolide A lost inhibitory activity (up to 20-fold) against the mutant T139Y, T139Q, T139K, and T139I enzymes. All of the mutant enzymes retained marked susceptibility toward the other NNRTIs, including nevirapine, delavirdine, efavirenz, thiocarboxanilide UC-781, quinoxaline GW867420X, TSAO [[2',5'-bis-O-(tert-butyldimethylsilyl)-beta-D-ribofuranosyl]-3'-spiro-5 ''-(4 ''-amino-1 '',2 ''-oxathiole-2 '',2 ''-dioxide)]derivatives, and the nucleoside inhibitor, ddGTP. The fact that the T139I RT 1) proved to be resistant to (+)-calanolide A, 2) represents a catalytically efficient enzyme, and 3) requires only a single transition point mutation (ACA -> ATA) in codon 139 seems to explain why mutant T139I RT virus strains, but not virus strains containing other amino acid changes at this position, predominantly emerge in cell cultures under (+)-calanolide A pressure.
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页码:652 / 659
页数:8
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