Specific substrates for spectrophotometric determination of penicillin acylase activity.
Penicillin acylase substrates suitable for colorimetric determination of the enzyme activity have been tested in this study. The kinetic parameters (Km and kcat) have been elucidated for the following nine substrates: six phenylacetic acid derivatives (p-nitroanilide, p-nitrophenyl ester, p-nitro-m-carboxyanilide, p-nitro-o-carboxyanilide, p-nitro-o-hydroxyanilide, m-nitro-p-carboxyanilide), two D-phenylglycine derivatives (p-nitroanilide, p-nitro-m-carboxyanilide), and also p-nitrophenyl ester of acetic acid (p-nitrophenyl acetate). With the exception of p-nitrophenyl acetate, all the compounds studied are highly specific chromogenic substrates for penicillin acylase, but their reactivity is very variable and kcat/Km values are in a range from 0.8.10(4) to 5.10(6) M(-1).sec(-1).
Youshko
M I
MI
Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, 119899, Russia.
Shamolina
T A
TA
Guranda
D F
DF
Synev
A V
AV
Svedas
V K
VK
D000813:Anilides / Q000378:metabolism; D020134:Catalytic Domain; D002863:Chromogenic Compounds; D003124:Colorimetry; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D010399:Penicillin Amidase / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D013053:Spectrophotometry; D013379:Substrate Specificity
D000813:Anilides; D002863:Chromogenic Compounds; D010399:Penicillin Amidase