Affinities of penicillins and cephalosporins for the penicillin-binding proteins of Escherichia coli K-12 and their antibacterial activity

Curtis, Nigel A. C.; Orr, David C.; Ross, Gayle; Boulton, M. G.

doi:10.1128/aac.16.5.533

Cited by 168 publications

(140 citation statements)

References 22 publications

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“…To identify variants of PBP1b that no longer require LpoB (designated PBP1b*), we used the cefsulodinhypersensitivity phenotype of mutants inactivated for PBP1b (ΔponB) or LpoB (ΔlpoB) (9,10). Cefsulodin is a β-lactam that preferentially targets PBP1a and PBP1b in E. coli (15). We reasoned that the cefsulodin hypersensitivity of ΔlpoB mutants likely results from a defect in PBP1b activity and that ponB* alleles encoding PBP1b* variants might be identified by a selection for increased cefsulodin resistance in a LpoB − strain background.…”

Section: Resultsmentioning

confidence: 99%

Cofactor bypass variants reveal a conformational control mechanism governing cell wall polymerase activity

Markovski

Bohrhunter

Lupoli

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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To fortify their cytoplasmic membrane and protect it from osmotic rupture, most bacteria surround themselves with a peptidoglycan (PG) exoskeleton synthesized by the penicillin-binding proteins (PBPs). As their name implies, these proteins are the targets of penicillin and related antibiotics. We and others have shown that the PG synthases PBP1b and PBP1a of Escherichia coli require the outer membrane lipoproteins LpoA and LpoB, respectively, for their in vivo function. Although it has been demonstrated that LpoB activates the PG polymerization activity of PBP1b in vitro, the mechanism of activation and its physiological relevance have remained unclear. We therefore selected for variants of PBP1b (PBP1b*) that bypass the LpoB requirement for in vivo function, reasoning that they would shed light on LpoB function and its activation mechanism. Several of these PBP1b variants were isolated and displayed elevated polymerization activity in vitro, indicating that the activation of glycan polymer growth is indeed one of the relevant functions of LpoB in vivo. Moreover, the location of amino acid substitutions causing the bypass phenotype on the PBP1b structure support a model in which polymerization activation proceeds via the induction of a conformational change in PBP1b initiated by LpoB binding to its UB2H domain, followed by its transmission to the glycosyl transferase active site. Finally, phenotypic analysis of strains carrying a PBP1b* variant revealed that the PBP1b-LpoB complex is most likely not providing an important physical link between the inner and outer membranes at the division site, as has been previously proposed.T he peptidoglycan (PG) layer forms a protective shell that surrounds the cytoplasmic membrane of bacteria to prevent osmotic rupture and provide cells with their characteristic shape (1). This complex macromolecule is composed of glycan strands crosslinked to one another by attached peptide chains to form the exoskeletal matrix. Because of its essentiality and uniqueness to bacteria, the PG layer is an important therapeutic target. Many antibiotics in our current arsenal block PG assembly, with penicillin and related beta-lactam drugs being the most wellknown and widely used. These molecules target major PG synthase enzymes called penicillin-binding proteins (PBPs) (1).The PBPs function in the final stage of the three-part pathway for PG biogenesis (1). Precursor synthesis begins in the cytoplasm with the production of the activated sugar molecules uridine diphosphate (UDP)-N-acetylmuramic acid pentapeptide (UDP-MurNAc-pep 5 ) and UDP-N-acetylglucosamine (UDPGlcNAc). In the second, membrane-associated phase, UDPMurNAc-pep 5 is converted to the precursor lipid-I by MraY, which transfers phospho-MurNAc-pep to the lipid carrier undecaprenol-phosphate (Und-P). Lipid-II is formed by MurG via the addition of GlcNAc to lipid-I from UDP-GlcNAc. This final precursor contains the basic monomeric unit of PG, the disaccharide-pentapeptide. After its production, lipid-II must be flipped by MurJ (2, 3) ...

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Section: Resultsmentioning

confidence: 99%

Cofactor bypass variants reveal a conformational control mechanism governing cell wall polymerase activity

Markovski

Bohrhunter

Lupoli

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…PBPs 1a and 1b are required for peptidoglycan synthesis, and inhibition of these enzymes, in the absence of peptidoglycan hydrolase inhibition, is thought to explain spheroplast formation [75]. For β-lactams which have a higher affinity for E. coli PBPs 1a and 1b than PBP2 or PBP3, some examples being amoxycillin, cephaloridine, and cefsulodin, spheroplast formation is the primary morphological response to antibiotic treatment [76].…”

Section: Spheroplast and Protoplast Formationmentioning

confidence: 99%

“…In Escherichia coli, β-lactam-induced spheroplast formation is due to inhibition of penicillin binding proteins 1a and 1b (PBPs 1a and 1b) [75][76][77]. PBPs 1a and 1b are required for peptidoglycan synthesis, and inhibition of these enzymes, in the absence of peptidoglycan hydrolase inhibition, is thought to explain spheroplast formation [75].…”

Section: Spheroplast and Protoplast Formationmentioning

confidence: 99%

Morphological and ultrastructural changes in bacterial cells as an indicator of antibacterial mechanism of action

Cushnie

O’Driscoll

Lamb

2016

Cell. Mol. Life Sci.

149

124

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“…We considered, therefore, the effects of 3 other ␤-lactams (Fig. S4): ampicillin, which inhibits multiple targets (PBP1a, PBP1b, and PBP3), cefsulodin, which is selective for PBP1a and PBP1b (25), and piperacillin, another selective inhibitor of PBP3. We first measured the protection of sulA-induced cells against these additional drugs and confirmed that inhibition of divisome assembly protects very powerfully against fast lysis promoted by cephalexin (Fig.…”

Section: Sensitivity To ␤-Lactams Depends On the State Of Divisome Asmentioning

confidence: 99%

Rapid β-lactam-induced lysis requires successful assembly of the cell division machinery

Chung

Yao

Goehring³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

108

116

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␤-lactam antibiotics inhibit penicillin binding proteins (PBPs) involved in peptidoglycan synthesis. Although inhibition of peptidoglycan biosynthesis is generally thought to induce cell lysis, the pattern and mechanism of cell lysis can vary substantially. ␤-lactams that inhibit FtsI, the only division specific PBP, block cell division and result in growth as filaments. These filaments ultimately lyse through a poorly understood mechanism. Here we find that one such ␤-lactam, cephalexin, can, under certain conditions, lead instead to rapid lysis at nascent division sites through a process that requires the complete and ordered assembly of the divisome, the essential machinery involved in cell division. We propose that this assembly process (in which the localization of cell wall hydrolases depends on properly targeted FtsN, which in turn depends on the presence of FtsI) ensures that the biosynthetic machinery to form new septa is in place before the machinery to degrade septated daughter cells is enabled. ␤-lactams that target FtsI subvert this mechanism by inhibiting FtsI without perturbing the normal assembly of the cell division machinery and the consequent activation of cell wall hydrolases. One seemingly paradoxical implication of our results is that ␤-lactam therapy may be improved by promoting active cell division.amidase ͉ cell wall hydrolase ͉ ftsN ͉ penicillin-binding proteins ͉ peptidoglycan

show abstract

Affinities of penicillins and cephalosporins for the penicillin-binding proteins of Escherichia coli K-12 and their antibacterial activity

Cited by 168 publications

References 22 publications

Cofactor bypass variants reveal a conformational control mechanism governing cell wall polymerase activity

Cofactor bypass variants reveal a conformational control mechanism governing cell wall polymerase activity

Morphological and ultrastructural changes in bacterial cells as an indicator of antibacterial mechanism of action

Rapid β-lactam-induced lysis requires successful assembly of the cell division machinery

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