Role of Inhibition of Penicillin Binding Proteins and Cell Wall Cross-Linking by Beta-Lactam Antibiotics in Low- and High-Level Methicillin Resistance of Staphylococcus aureus

Chemotherapy ◽  
1999 ◽  
Vol 45 (1) ◽  
pp. 37-47 ◽  
Author(s):  
Yasuyuki Higashi ◽  
Akiko Wakabayashi ◽  
Yoshimi Matsumoto ◽  
Yuji Watanabe ◽  
Akira Ohno
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Binding Proteins ◽  
Methicillin Resistance ◽  
Cross Linking ◽  
Beta Lactam ◽  
Penicillin Binding Proteins ◽  
Beta Lactam Antibiotics ◽  
High Level

scholarly journals Penicillin-binding proteins 2b and 2x of Streptococcus pneumoniae are primary resistance determinants for different classes of beta-lactam antibiotics.

1996 ◽  
Vol 40 (4) ◽  
pp. 829-834 ◽  
Author(s):  
T Grebe ◽  
R Hakenbeck
Keyword(s):  
Streptococcus Pneumoniae ◽  
Clinical Isolate ◽  
Binding Proteins ◽  
Point Mutations ◽  
Clinical Isolates ◽  
Single Amino Acid ◽  
Beta Lactam ◽  
Penicillin Binding Proteins ◽  
Beta Lactam Antibiotics ◽  
High Level

High-level resistance to beta-lactam antibiotics in Streptococcus pneumoniae is mediated by successive alterations in essential penicillin-binding proteins (PBPs). In the present work, single amino acid changes in S. pneumoniae PBP 2x and PBP 2b that result in reduced affinity for the antibiotic and that confer first-level beta-lactam resistance are defined. Point mutations in the PBP genes were generated by PCR-derived mutagenesis. Those conferring maximal resistance to either cefotaxime (pbp2x) or piperacillin (pbp2b) were obtained after transformation of the susceptible laboratory strain R6 with the PCR-amplified PBP genes and selection on agar with various concentrations of the antibiotic. In the case of PBP 2x, transformants for which the cefotaxime MIC was 0.16 microgram/ml contained the substitution of a Thr for an Ala at position 550 (Thr550-->Ala), close to the PBP homology box Lys547SerGly, a mutation frequently observed in laboratory mutants and in a high-level cefotaxime-resistant clinical isolate as well. After further selection, transformants resisting 0.3 microgram of cefotaxime per ml were obtained; they contained the substitution Gly550 as the result of two mutations in the same codon. In PBP 2b, Thr446-->Ala, adjacent to another homology box Ser443SerAsn, was the mutation selected with piperacillin. This substitution has been described in all clinical isolates with a low-affinity PBP 2b but was distinct from point mutations found in laboratory mutants. Both pbp2b with the single mutation and a mosaic pbp2b of a clinical isolate conferred a twofold increase in piperacillin resistance. Attempts to select PBP 2b variants at higher piperacillin concentrations were unsuccessful. The mutated PBP 2b also markedly reduced the lytic response to piperacillin, suggesting that such a mutation is an important step in resistance development in clinical isolates.

scholarly journals Effects of inhibitors of protein synthesis on lysis of Escherichia coli induced by beta-lactam antibiotics.

1996 ◽  
Vol 40 (4) ◽  
pp. 899-903 ◽  
Author(s):  
D G Rodionov ◽  
E E Ishiguro
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Binding Proteins ◽  
Rapid Development ◽  
Stringent Response ◽  
Beta Lactam ◽  
Penicillin Binding Proteins ◽  
Beta Lactam Antibiotics ◽  
Inhibition Of Protein Synthesis

The role of protein synthesis in ampicillin-induced lysis of Escherichia coli was investigated. The inhibition of protein synthesis through amino acid deprivation resulted in the rapid development of ampicillin tolerance as a consequence of the stringent response, as previously reported. In contrast, inhibition of protein synthesis by use of ribosome inhibitors such as chloramphenicol did not readily confer ampicillin tolerance and, in fact, promoted the development of both stages of the ampicillin-induced lysis process, i.e., (i) an ampicillin-dependent stage which apparently involves the interaction of penicillin-binding proteins with ampicillin and (ii) an ampicillin-independent stage which may represent the events leading to the deregulation of peptidoglycan hydrolase activity. We propose that lysis was facilitated when protein synthesis was inhibited because the production of new penicillin-binding proteins to replace those which were ampicillin inhibited was prevented under these conditions.

scholarly journals Role of murE in the Expression of β-Lactam Antibiotic Resistance in Staphylococcus aureus

2004 ◽  
Vol 186 (6) ◽  
pp. 1705-1713 ◽  
Author(s):  
S. Gardete ◽  
A. M. Ludovice ◽  
R. G. Sobral ◽  
S. R. Filipe ◽  
H. de Lencastre ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Methicillin Resistance ◽  
Inducible Promoter ◽  
Northern Analysis ◽  
Penicillin Binding Protein ◽  
Drastic Reduction ◽  
Precursor Pool ◽  
Lactam Antibiotic

ABSTRACT It was shown earlier that Tn551 inserted into the C-terminal region of murE of parental methicillin-resistant Staphylococcus aureus strain COL causes a drastic reduction in methicillin resistance, accompanied by accumulation of UDP-MurNAc dipeptide in the cell wall precursor pool and incorporation of these abnormal muropeptides into the peptidoglycan of the mutant. Methicillin resistance was recovered in a suppressor mutant. The murE gene of the same strain was then put under the control of the isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible promoter P spac . Bacteria grown in the presence of suboptimal concentrations of IPTG accumulated UDP-MurNAc dipeptide in the cell wall precursor pool. Both growth rates and methicillin resistance levels (but not resistance to other antibiotics) were a function of the IPTG concentration. Northern analysis showed a gradual increase in the transcription of murE and also in the transcription of pbpB and mecA, parallel with the increasing concentrations of IPTG in the medium. A similar increase in the transcription of pbpB and mecA, the structural genes of penicillin-binding protein 2 (PBP2) and PBP2A, was also detected in the suppressor mutant. The expression of these two proteins, which are known to play critical roles in the mechanism of staphylococcal methicillin resistance, appears to be—directly or indirectly—under the control of the murE gene. Our data suggest that the drastic reduction of the methicillin MIC seen in the murE mutant may be caused by the insufficient cellular amounts of these two PBPs.

scholarly journals FmhA and FmhC of Staphylococcus aureus incorporate serine residues into peptidoglycan cross-bridges

2020 ◽  
Vol 295 (39) ◽  
pp. 13664-13676 ◽  
Author(s):  
Stephanie Willing ◽  
Emma Dyer ◽  
Olaf Schneewind ◽  
Dominique Missiakas
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Binding Proteins ◽  
Penicillin Binding Proteins ◽  
Daughter Cells ◽  
Cross Bridge ◽  
Resistant Strains ◽  
The Cross ◽  
Cross Bridges ◽  
Adjacent Wall

Staphylococcal peptidoglycan is characterized by pentaglycine cross-bridges that are cross-linked between adjacent wall peptides by penicillin-binding proteins to confer robustness and flexibility. In Staphylococcus aureus, pentaglycine cross-bridges are synthesized by three proteins: FemX adds the first glycine, and the homodimers FemA and FemB sequentially add two Gly-Gly dipeptides. Occasionally, serine residues are also incorporated into the cross-bridges by enzymes that have heretofore not been identified. Here, we show that the FemA/FemB homologues FmhA and FmhC pair with FemA and FemB to incorporate Gly-Ser dipeptides into cross-bridges and to confer resistance to lysostaphin, a secreted bacteriocin that cleaves the pentaglycine cross-bridge. FmhA incorporates serine residues at positions 3 and 5 of the cross-bridge. In contrast, FmhC incorporates a single serine at position 5. Serine incorporation also lowers resistance toward oxacillin, an antibiotic that targets penicillin-binding proteins, in both methicillin-sensitive and methicillin-resistant strains of S. aureus. FmhC is encoded by a gene immediately adjacent to lytN, which specifies a hydrolase that cleaves the bond between the fifth glycine of cross-bridges and the alanine of the adjacent stem peptide. In this manner, LytN facilitates the separation of daughter cells. Cell wall damage induced upon lytN overexpression can be alleviated by overexpression of fmhC. Together, these observations suggest that FmhA and FmhC generate peptidoglycan cross-bridges with unique serine patterns that provide protection from endogenous murein hydrolases governing cell division and from bacteriocins produced by microbial competitors.

scholarly journals Role of the msaABCR Operon in Cell Wall Biosynthesis, Autolysis, Integrity, and Antibiotic Resistance in Staphylococcus aureus

2019 ◽  
Vol 63 (10) ◽  
Author(s):  
Bibek G C ◽  
Gyan S. Sahukhal ◽  
Mohamed O. Elasri
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Antibiotic Resistance ◽  
Teichoic Acid ◽  
Cross Linking ◽  
Wall Defect ◽  
Content Type ◽  
Mrsa Strains ◽  
Mutant Cells

ABSTRACT Staphylococcus aureus is an important human pathogen in both community and health care settings. One of the challenges with S. aureus as a pathogen is its acquisition of antibiotic resistance. Previously, we showed that deletion of the msaABCR operon reduces cell wall thickness, resulting in decreased resistance to vancomycin in vancomycin-intermediate S. aureus (VISA). In this study, we investigated the nature of the cell wall defect in the msaABCR operon mutant in the Mu50 (VISA) and USA300 LAC methicillin-resistant Staphylococcus aureus (MRSA) strains. Results showed that msaABCR mutant cells had decreased cross-linking in both strains. This defect is typically due to increased murein hydrolase activity and/or nonspecific processing of murein hydrolases mediated by increased protease activity in mutant cells. The defect was enhanced by a decrease in teichoic acid content in the msaABCR mutant. Therefore, we propose that deletion of the msaABCR operon results in decreased peptidoglycan cross-linking, leading to increased susceptibility toward cell wall-targeting antibiotics, such as β-lactams and vancomycin. Moreover, we also observed significantly downregulated transcription of early cell wall-synthesizing genes, supporting the finding that msaABCR mutant cells have decreased peptidoglycan synthesis. More specifically, the msaABCR mutant in the USA300 LAC strain (MRSA) showed significantly reduced expression of the murA gene, whereas the msaABCR mutant in the Mu50 strain (VISA) showed significantly reduced expression of glmU, murA, and murD. Thus, we conclude that the msaABCR operon controls the balance between cell wall synthesis and cell wall hydrolysis, which is required for maintaining a robust cell wall and acquiring resistance to cell wall-targeting antibiotics, such as vancomycin and the β-lactams.

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