MECHANISM OF PENICILLIN ACTION - PENICILLIN AND SUBSTRATE BIND COVALENTLY TO THE SAME ACTIVE-SITE SERINE IN 2 BACTERIAL D-ALANINE CARBOXYPEPTIDASES

被引:112
作者
YOCUM, RR
WAXMAN, DJ
RASMUSSEN, JR
STROMINGER, JL
机构
关键词
D O I
10.1073/pnas.76.6.2730
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
it has been hypothesized that penicillin acts as a structural analog of the acyl-D-alanyl-D-alanine terminus of nascent bacterial cell wall and that it consequently binds to and acylates the active site of the enzyme(s) that crosslinks the cell wall to form an inactive penicilloyl enzyme [Tipper, D.J. & Strominger, J.L. (1965) Proc. Natl. Acad. Sci. USA 64, 1133-1138]. This study directly proves that penicillin acylates the active site of two penicillin-sensitive enzymes, D-alanine carboxypeptidases from Bacillus stearothermophilus and Bacillus subtilis. Active site peptides were degenerated by chemical or enzymatic cleavage of these carboxypeptidases after covalently labeling with [14C]penicillin G or after trapping an acylenzyme intermediate derived from the depsipeptide substrate, [14C]diacetyl-L-lysyl-D-alanyl-D-lactate. The aminoacid sequences of the penicillin- and substrate-labeled peptides were identical. Both penicillin and substrate were covalently bound via an ester linkage to the same active site residue, a serine at position 36 of the B. stearothermophilus carboxypeptidase and the corresponding serine in the B. subtilis carboxypeptidase. The two D-alanine carboxypeptidases showed significant homology around the active site. Moreover, homology between these two enzymes and four β-lactamases of known sequence suggests that these two groups of enzymes are evolutionally related.
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页码:2730 / 2734
页数:5
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