A Role in vivo for Penicillin‐Binding Protein‐4 of Staphylococcus aureus

Abstract

The degree of cross-linking of the peptidoglycan of Staphylococcus aureus H and mutants lacking penicillinbinding proteins 1 and 4 was studied. No major changes were observed in organisms lacking protein 1 whereas loss of protein 4 was accompanied by a marked reduction in the degree of cross-linking and the absence of a membrane-bound 'model' transpeptidase activity. A similar effect was achieved when cultures of the staphylococci were treated with the P-lactam antibiotic cefoxitin. At low concentrations (0.05 pg m1-l) cefoxitin shows highest affinity for protein 4 to which it appears to bind irreversibly. Treatment of the mutant lacking protein 4 with this concentration of the antibiotic did not affect the degree of cross-linkage.The possibility that the decrease in cross-linkage was a consequence of DD-carboxypeptidase activity on peptidoglycan precursors was investigated. Although both S. uureus H and the mutants possessed such activity it was insensitive to benzylpenicillin and cefoxitin and the role of this enzyme(s) in peptidoglycan biosynthesis remains unknown.We conclude that in vivo protein 4 acts as a transpeptidase involved in the secondary cross-linking of peptido-,glycan and this activity is necessary to achieve the high degree of cross-linkage observed in the peptidoglycan of staphylococci.The penicillin-binding proteins of both gram-positive and gram-negative bacteria have received considerable attention in recent years [I -31. These investigations have resulted in the general conclusion that the interaction of B-lactams with one or more of the binding proteins is responsible for the lethal effects of these antibiotics. However, only in Escherichia coli has the role of individual penicillin-binding proteins been investigated in detail. In this organism distinct penicillinbinding proteins have been implicated in the determination of cell shape [4,5] ; cell division [6] and elongation [4,5,7,8]. With the exception of penicillin-binding protein 2, all have now been isolated and purified [9-141 and in the case of protein 1A [I21 and 1B [9,10,13] shown to catalyse in vitro transglycosylation and transpeptidation reactions using lipid intermediates as the peptidoglycan precursors. The lower molecular weight proteins (4, 5 and 6) catalyse DD-carboxypeptidase and in the case of proteins 5 and 6 'model' transpeptidase activity [I 1,141. Mutants lacking these proteins (4 = ducB 5 = ducA) grow normally under various conditions and it was concluded these proteins were 'non-essential' Similar studies on gram-positive bacteria have tended to concentrate on Staphylococcus aureus and various Bacillus spp. (For review, see [2].) In the bacilli, four to six distinct penicillin-binding proteins have been described in which the one of lowest molecular weight (protein 5 in Bacillus subtilis) has DD-carboxypeptidase and 'model' transpeptidase activity [I 9,201 Enzyme. DD-Carboxypeptidase or muramoyl-pentapeptide carboxypeptidase (EC 3.4.17.8).H. A. Chase, unpublished observations). A similar situation exists in S. uur...

Multiplex PCR for the Detection of Genes Encoding Aminoglycoside Modifying Enzymes and Methicillin Resistance among Staphylococcus Species

“…However, 11% (10/92) of the strains were discordant, and most of these were CNS (7 strains of OX r /mecAand 2 strains of OX s /mecA + ). In the case of OX r /mecA -strains, non-PBP 2a-dependent mechanisms such as hyperproduction of -lactamase and alteration of PBP types may participate in the expression of resistance (20,21). We performed nitrocefin disk test (Becton Dickinson, Sparks, U.S.A.) for the 5 strain out of 7 discordant OX r /mecA -strain and 3 strain showed positive results.…”

Occurrence of a beta-lactam-inducible penicillin-binding protein in methicillin-resistant staphylococci

“…However, PBP-1 and PBP-4 are not essential targets since a mutant lacking these PBPs is viable (37). Thus, it is highly probable that PBP-2 or PBP-3 is the lethal target.…”

Increased amounts of a novel penicillin-binding protein in a strain of methicillin-resistant Staphylococcus aureus exposed to nafcillin.

“…Bacterial species may synthesize several PBPs, each serving a different function (1)(2)(3). Because to some extent one PBP can substitute for the function of another PBP, probably not all are essential for cell growth (4,5).…”