Selective and facile assay of human immunodeficiency virus protease activity by a novel fluorogenic reaction.
A highly selective and facile assay of human immunodeficiency virus protease (HIV-PR) has been required for the screening of medicinal inhibitors and also for classifying the subtypes of HIV in the therapeutic treatment of acquired immune deficiency syndrome (AIDS). This article describes a novel assay method of HIV-PR based on the selective fluorogenic reaction of peptides. A peptide fragment generated from a substrate by the enzymatic digestion with HIV-PR could be selectively quantified by the spectrofluorometric detection after the fluorogenic reaction with catechol in the presence of sodium periodate and sodium borate (pH 7.0). This assay system uses an N-terminal acetyl peptide as the substrate and crude extracts from Escherichia coli expressing recombinant HIV-PR. The activity obtained by the proposed assay correlated with that obtained by a conventional HIV-PR assay based on fluorescence resonance energy transfer detection.
Yu
Zhiqiang
Z
Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8521, Japan.
Kabashima
Tsutomu
T
Tang
Chenhong
C
Shibata
Takayuki
T
Kitazato
Kaio
K
Kobayashi
Nobuyuki
N
Lee
Myung Koo
MK
Kai
Masaaki
M
D016333:HIV Protease / Q000032:analysis* / Q000378:metabolism; D017320:HIV Protease Inhibitors / Q000494:pharmacology; D015497:HIV-1 / Q000201:enzymology; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence
D017320:HIV Protease Inhibitors; D016333:HIV Protease