Open AccessPurification and Characterization of PBP4a, a New Low-Molecular-Weight Penicillin-Binding Protein from Bacillus subtilis
Author(s) -
Colette Duez,
Marc Vanhove,
Xavier Gallet,
Fabrice Bouillenne,
Jean Denis Docquier,
Alain Brans,
JeanMarie Frère
Publication year - 2001
Publication title -
journal of bacteriology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.652
H-Index - 246
eISSN - 1067-8832
pISSN - 0021-9193
DOI - 10.1128/jb.183.5.1595-1599.2001
Subject(s) - bacillus subtilis , biology , escherichia coli , penicillin binding proteins , biochemistry , cephaloridine , benzylpenicillin , microbiology and biotechnology , serine , enzyme , carboxypeptidase , penicillin , acylation , strain (injury) , bacteria , antibiotics , cephalosporin , genetics , catalysis , gene , anatomy
Penicillin-binding protein 4a (PBP4a) from Bacillus subtilis was overproduced and purified to homogeneity. It clearly exhibits DD-carboxypeptidase and thiolesterase activities in vitro. Although highly isologous to the Actinomadura sp. strain R39 DD-peptidase (B. Granier, C. Duez, S. Lepage, S. Englebert, J. Dusart, O. Dideberg, J. van Beeumen, J. M. Frère, and J. M. Ghuysen, Biochem. J. 282:781-788, 1992), which is rapidly inactivated by many beta-lactams, PBP4a is only moderately sensitive to these compounds. The second-order rate constant (k(2)/K) for the acylation of the essential serine by benzylpenicillin is 300,000 M(-1) s(-1) for the Actinomadura sp. strain R39 peptidase, 1,400 M(-1) s(-1) for B. subtilis PBP4a, and 7,000 M(-1) s(-1) for Escherichia coli PBP4, the third member of this class of PBPs. Cephaloridine, however, efficiently inactivates PBP4a (k(2)/K = 46,000 M(-1) s(-1)). PBP4a is also much more thermostable than the R39 enzyme.