Affinities of β-Lactams for Penicillin Binding Proteins of
            <i>Chlamydia trachomatis</i>
            and Their Antichlamydial Activities

Storey, Christopher; Chopra, Ian

doi:10.1128/aac.45.1.303-305.2001

Cited by 25 publications

(24 citation statements)

References 22 publications

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“…We found that PBP1a pathway mutants are highly susceptible to this cephalosporin antibiotic, suggesting that it inhibits PBP1b exclusively. Though it is well established that beta-lactam and cephalosporin antibiotics have differing affinities for different PBPs (28)(29)(30), such a stark difference is unusual. Similarly, we found that penicillin G and ticarcillin, broad-spectrum inhibitors of PBPs, were effective against PBP1a pathway mutants at lower concentrations than those that are effective against the wild type or the PBP1b pathway mutants.…”

Section: Discussionmentioning

confidence: 99%

Differential Requirement for PBP1a and PBP1b in In Vivo and In Vitro Fitness of Vibrio cholerae

et al. 2014

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We investigated the roles of the Vibrio cholerae high-molecular-weight bifunctional penicillin binding proteins, PBP1a and PBP1b, in the fitness of this enteric pathogen. Using a screen for synthetic lethality, we found that the V. cholerae PBP1a and PBP1b proteins, like their Escherichia coli homologues, are each essential in the absence of the other and in the absence of the other's putative activator, the outer membrane lipoproteins LpoA and LpoB, respectively. Comparative analyses of V. cholerae mutants suggest that PBP1a/LpoA of V. cholerae play a more prominent role in generating and/or maintaining the pathogen's cell wall than PBP1b/LpoB. V. cholerae lacking PBP1b or LpoB exhibited wild-type growth under all conditions tested. In contrast, V. cholerae lacking PBP1a or LpoA exhibited growth deficiencies in minimal medium, in the presence of deoxycholate and bile, and in competition assays with wild-type cells both in vitro and in the infant mouse small intestine. PBP1a pathway mutants are particularly impaired in stationary phase, which renders them sensitive to a product(s) present in supernatants from stationary-phase wild-type cells. The marked competitive defect of the PBP1a pathway mutants in vivo was largely absent when exponential-phase cells rather than stationary-phase cells were used to inoculate suckling mice. Thus, at least for V. cholerae PBP1a pathway mutants, the growth phase of the inoculum is a key modulator of infectivity.T he main component of the bacterial cell wall, peptidoglycan (PG), is an intricate mesh of polysaccharide chains crosslinked by short peptide bridges. The periplasmic assembly of this complex polymer from [N-acetylglucosamine-N-acetylmuramic acid]pentapeptide subunits is facilitated by extracytoplasmic enzymes known as penicillin binding proteins (PBPs). These enzymes carry out two main reactions: transglycosylation (i.e., polymerization of glycan subunits) and transpeptidation (i.e., cross-linking the peptide side chains between polymerized glycan strands). In Escherichia coli-the focus of most studies of Gram-negative cell wall synthesis-two principal high-molecular-weight (HMW) PBPs with both transglycosylase and transpeptidase activity play a pivotal role in PG synthesis. E. coli PBP1a and PBP1b appear to be largely interchangeable, and mutants lacking one of the two proteins have at most mild phenotypes under standard growth conditions (1-3). However, cells lacking both proteins are not viable, and PBP1a and PBP1b are termed synthetically lethal. The activity of each PBP1 enzyme is strictly dependent on the presence of a specific outer membrane activator, either LpoA (for PBP1a) or LpoB (for PBP1b), and mutations in either lpo locus are consequently also synthetically lethal with a mutation of the noncognate pbp1 locus (4, 5). It has been proposed that PBP1a may contribute preferentially to cell elongation whereas PBP1b may play a more prominent role in cell division (3, 6); however, the viability of the individual mutants clearly demonstrates that each enzyme can ...

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Section: Discussionmentioning

confidence: 99%

Differential Requirement for PBP1a and PBP1b in In Vivo and In Vitro Fitness of Vibrio cholerae

et al. 2014

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“…In the periplasm, penicillin-binding proteins cross-link the subunits, giving rise to PG layers that expand the size and volume of RBs and facilitate cell division. Further support for this model includes the expression of pbp2 mRNA transcripts at 6 to 8 hpi (data not shown) and the effectiveness of cell wall antibiotics, which interfere with RB development and cell division (19,21,35).…”

Section: Discussionmentioning

confidence: 99%

“…Recently, it was shown that mecillinam specifically binds to the chlamydial PBP2 homologue in vitro and inhibits chlamydial development in vivo, although the effect of mecillinam on cell morphology was not studied (35). It is not known whether the chlamydial PBPs have transpeptidase activity, as these enzymes have to date been characterized only by their ability to bind radiolabeled ␤-lactams (2,35). It is possible that the roles of PBP2 and PBP3 in the formation of chlamydial PG differ from their roles in E. coli.…”

Section: Discussionmentioning

confidence: 99%

In Vitro and In Vivo Functional Activity ofChlamydiaMurA, a UDP-N-Acetylglucosamine Enolpyruvyl Transferase Involved in Peptidoglycan Synthesis and Fosfomycin Resistance

2003

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Organisms of Chlamydia spp. are obligate intracellular, gram-negative bacteria with a dimorphic developmental cycle that takes place entirely within a membrane-bound vacuole termed an inclusion. The chlamydial anomaly refers to the fact that cell wall-active antibiotics inhibit Chlamydia growth and peptidoglycan (PG) synthesis genes are present in the genome, yet there is no biochemical evidence for synthesis of PG. In this work, we undertook a genetics-based approach to reevaluate the chlamydial anomaly by characterizing MurA, a UDP-N-acetylglucosamine enolpyruvyl transferase that catalyzes the first committed step of PG synthesis. The murA gene from Chlamydia trachomatis serovar L2 was cloned and placed under the control of the arabinose-inducible, glucose-repressible ara promoter and transformed into Escherichia coli. After transduction of a lethal ⌬murA mutation into the strain, viability of the E. coli strain became dependent upon expression of the C. trachomatis murA. DNA sequence analysis of murA from C. trachomatis predicted a cysteine-toaspartate change in a key residue within the active site of MurA. In E. coli, the same mutation has previously been shown to cause resistance to fosfomycin, a potent antibiotic that specifically targets MurA. In vitro activity of the chlamydial MurA was resistant to high levels of fosfomycin. Growth of C. trachomatis was also resistant to fosfomycin. Moreover, fosfomycin resistance was imparted to the E. coli strain expressing the chlamydial murA. Conversion of C. trachomatis elementary bodies to reticulate bodies and cell division are correlated with expression of murA mRNA. mRNA from murB, the second enzymatic reaction in the PG pathway, was also detected during C. trachomatis infection. Our findings, as well as work from other groups, suggest that a functional PG pathway exists in Chlamydia spp. We propose that chlamydial PG is essential for progression through the developmental cycle as well as for cell division. Elucidating the existence of PG in Chlamydia spp. is of significance for the development of novel antibiotics targeting the chlamydial cell wall.

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“…They undergo binary fission as reticulate bodies within the chlamydial inclusion. They have the information for the synthesis of a lipid II precursor similar to that of E. coli (216). However, they are peptidoglycanless.…”

Section: Where Next?mentioning

confidence: 99%

Biochemistry and Comparative Genomics of SxxK Superfamily Acyltransferases Offer a Clue to the Mycobacterial Paradox: Presence of Penicillin-Susceptible Target Proteins versus Lack of Efficiency of Penicillin as Therapeutic Agent

Goffin

Ghuysen

2002

Microbiol Mol Biol Rev

179

213

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The bacterial acyltransferases of the SxxK superfamily vary enormously in sequence and function, with conservation of particular amino acid groups and all-α and α/β folds. They occur as independent entities (free-standing polypeptides) and as modules linked to other polypeptides (protein fusions). They can be classified into three groups. The group I SxxK d,d-acyltransferases are ubiquitous in the bacterial world. They invariably bear the motifs SxxK, SxN(D), and KT(S)G. Anchored in the plasma membrane with the bulk of the polypeptide chain exposed on the outer face of it, they are implicated in the synthesis of wall peptidoglycans of the most frequently encountered (4→3) type. They are inactivated by penicillin and other β-lactam antibiotics acting as suicide carbonyl donors in the form of penicillin-binding proteins (PBPs). They are components of a morphogenetic apparatus which, as a whole, controls multiple parameters such as shape and size and allows the bacterial cells to enlarge and duplicate their particular pattern. Class A PBP fusions comprise a glycosyltransferase module fused to an SxxK acyltransferase of class A. Class B PBP fusions comprise a linker, i.e., protein recognition, module fused to an SxxK acyltransferase of class B. They ensure the remodeling of the (4→3) peptidoglycans in a cell cycle-dependent manner. The free-standing PBPs hydrolyze d,d peptide bonds. The group II SxxK acyltransferases frequently have a partially modified bar code, but the SxxK motif is invariant. They react with penicillin in various ways and illustrate the great plasticity of the catalytic centers. The secreted free-standing PBPs, the serine β-lactamases, and the penicillin sensors of several penicillin sensory transducers help the d,d-acyltransferases of group I escape penicillin action. The group III SxxK acyltransferases are indistinguishable from the PBP fusion proteins of group I in motifs and membrane topology, but they resist penicillin. They are referred to as Penr protein fusions. Plausible hypotheses are put forward on the roles that the Penr protein fusions, acting as l,d-acyltransferases, may play in the (3→3) peptidoglycan-synthesizing molecular machines. Shifting the wall peptidoglycan from the (4→3) type to the (3→3) type could help Mycobacterium tuberculosis and Mycobacterium leprae survive by making them penicillin resistant

show abstract

Affinities of β-Lactams for Penicillin Binding Proteins of Chlamydia trachomatis and Their Antichlamydial Activities

Cited by 25 publications

References 22 publications

Differential Requirement for PBP1a and PBP1b in In Vivo and In Vitro Fitness of Vibrio cholerae

Differential Requirement for PBP1a and PBP1b in In Vivo and In Vitro Fitness of Vibrio cholerae

In Vitro and In Vivo Functional Activity ofChlamydiaMurA, a UDP-N-Acetylglucosamine Enolpyruvyl Transferase Involved in Peptidoglycan Synthesis and Fosfomycin Resistance

Biochemistry and Comparative Genomics of SxxK Superfamily Acyltransferases Offer a Clue to the Mycobacterial Paradox: Presence of Penicillin-Susceptible Target Proteins versus Lack of Efficiency of Penicillin as Therapeutic Agent

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