ABSTRACT
Penicillin-binding proteins (PBPs) include transpeptidases, carboxypeptidases, and endopeptidases for biosynthesis of peptidoglycans in the cell wall to maintain bacterial morphology and survival in the environment. Streptococcus pneumoniae expresses six PBPs, but their enzymatic kinetic characteristics and inhibitory effects on different ß-lactam antibiotics remain poorly understood. In this study, all the six recombinant PBPs of S. pneumoniae displayed transpeptidase activity with different substrate affinities (Km = 1.56-9.11 mM) in a concentration-dependent manner, and rPBP3 showed a greater catalytic efficiency (Kcat = 2.38 s-1) than the other rPBPs (Kcat = 3.20-7.49 × 10-2 s-1). However, only rPBP3 was identified as a carboxypeptidase (Km = 8.57 mM and Kcat = 2.57 s-1). None of the rPBPs exhibited endopeptidase activity. Penicillin and cefotaxime inhibited the transpeptidase and carboxypeptidase activity of all the rPBPs but imipenem did not inhibited the enzymatic activities of rPBP3. Except for the lack of binding of imipenem to rPBP3, penicillin, cefotaxime, and imipenem bound to all the other rPBPs (KD = 3.71-9.35 × 10-4 M). Sublethal concentrations of penicillin, cefotaxime, and imipenem induced a decrease of pneumococcal pbps-mRNA levels (p < 0.05). These results indicated that all six PBPs of S. pneumoniae are transpeptidases, while only PBP3 is a carboxypeptidase. Imipenem has no inhibitory effect on pneumococcal PBP3. The pneumococcal genes for encoding endopeptidases remain to be determined.
