In addition to pbpA, encoding PBP2, other different genes such as rodA ( 59, 60) and mreBCD ( 33, 54, 60) are required for elongation in E. The composition of the multiprotein complex needed for cell elongation is not yet well documented, but it is thought to be structurally analogous to the divisome ( 30). These PBPs are part of a multiprotein-synthesizing machinery, designated the divisome ( 6, 21, 42, 54), whose assembly has been studied by different approaches ( 16, 31, 38, 45, 63). coli ( 38, 63, 65) or PBP2b and PBP1 of Bacillus subtilis ( 54) localize in the area of developing septa and of completed poles. In addition, two protein complexes containing different PBPs could be isolated from Haemophilus influenzae and Bacillus subtilis cells ( 1, 58).Ĭonsistent with their function, PBP3 of E. coli class B PBP2 and PBP3 either by β-lactam antibiotics or by mutagenesis shows that the first is specifically required for cell elongation and that the second is required for cell division ( 4, 59). Class B PBPs possess three conserved amino acid motifs in their N-terminal modules that have no detectable enzymatic activity but seem, however, to be essential for cell morphogenesis ( 26).ĭifferent observations suggest that rod-shaped bacteria possess two morphogenetic systems, one responsible for cell elongation and the other responsible for cytokinesis ( 50, 52). Class A PBPs have an N-terminal module acting as a glycosyltransferase and are characterized by the presence of five typical conserved amino acid motifs. They are further subdivided into two classes. In contrast, the high-molecular-mass PBPs are essential bimodular enzymes that bind penicillins on their C-terminal module. The low-molecular-mass proteins are essentially monomodular dd-peptidases that have been shown to play a morphogenetic role in Escherichia coli as well as in Streptococcus pneumoniae cells ( 11, 12, 41, 43, 66). The PBPs are separated into two large groups on the basis of their sizes. Generally, the PBPs lose their dd-peptidase activity by forming a long-lived covalent bond with penicillins ( 26). These peptidases, commonly designated penicillin-binding proteins (PBPs), are serine-active enzymes that belong to a large family of penicillin-recognizing proteins whose active sites contain three conserved motifs. The P 197-N 209 segment (site E) in one of these domains seemed to be particularly important, as single and double mutations reduced or almost completely abolished, respectively, the action of PBP5.Ĭell morphology in eubacteria is determined mainly by the peptidoglycan layer that is synthesized by a battery of cytoplasmic and membrane-bound enzymes, among which are the dd-peptidases ( 26, 42, 61). Mutations made in the two globular domains present in the N-terminal module indicated that they were needed for the suppletive activity. The T 167-L 184 region (designated site D) appeared to be an essential intramolecular site needed for the stability of the protein. Potential protein-protein interaction sites of the PBP5 N-terminal module were mutagenized by site-directed mutagenesis. Noninduced SL2 cells were unable to divide except when plasmid-borne pbp5 genes were present, provided that the PBP5 active site was functional. That substitution activity was analyzed in Enterococcus hirae SL2, a mutant whose pbp5 gene was interrupted by the nisRK genes and whose PBP3 synthesis was submitted to nisin induction. One important feature of PBP5 is its ability to substitute for most, if not all, penicillin-binding proteins when they are inhibited. The low susceptibility of enterococci to β-lactams is due to the activity of the low-affinity penicillin-binding protein 5 (PBP5).
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