Calcium-Dependent Complex Formation Between PBP2 and Lytic Transglycosylase SltB1 of<i>Pseudomonas aeruginosa</i>

Nikolaidis, Ioulia; Izoré, Thierry; Job, Viviana; Thielens, Nicole M.; Breukink, Eefjan; Dessen, Andréa

doi:10.1089/mdr.2012.0006

Cited by 26 publications

(36 citation statements)

References 29 publications

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“…SltB1 was previously reported to interact with PBP2 (19,20); therefore, we asked whether increased ␤-lactam resistance in the sltB1 deletion mutant was due to loss of its LT activity or the potential disruption of a PBP2-containing complex due to absence of the SltB1 protein. To address the second possibility, the sltB1 gene was replaced at its chromosomal locus (to ensure native expression levels) with mutant versions expressing one of two putative active-site mutants, one replacing the proposed catalytic Glu135 (21) with Ala and the other replacing Ser189, responsible for orientation of PG in the active site (22), with Ala.…”

Section: Resultsmentioning

confidence: 99%

“…Family designations for LTs are based on sequence motifs surrounding the catalytic regions; however, crystallographic studies with E. coli (32)(33)(34) and P. aeruginosa (20) showed that the catalytic domains of family 1 and family 3 LTs are structurally conserved despite their sequence dissimilarity, with a lysozyme fold and an essential catalytic Glu residue (35). One notable difference is the presence of an EF hand-type motif in the core domain of family 3 LTs.…”

Section: Figmentioning

confidence: 99%

“…One notable difference is the presence of an EF hand-type motif in the core domain of family 3 LTs. In SltB1, this motif contains a calciumbinding loop required for interaction with PBP2 and possibly other protein partners (20). Family 1 LTs have roles in conjugation and secretion systems, while family 3 LTs are involved in assembly of flagella and pili and in sporulation (36).…”

Section: Figmentioning

confidence: 99%

“…Furthermore, PG-active enzymes can be organized into multienzyme complexes (19,20,38), and loss of one component might destabilize those complexes, affecting PG metabolism. SltB1 interacts with PBP2 (19,20) but could have additional partners (20). Chromosomal expression of putative active-site mutants of SltB1 did not restore susceptibility, suggesting that increased ␤-lactam resistance in sltB1 mutants is due to the loss of SltB1 function, rather than disruption of protein interactions.…”

Section: Figmentioning

confidence: 99%

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Changes to Its Peptidoglycan-Remodeling Enzyme Repertoire Modulate β-Lactam Resistance in Pseudomonas aeruginosa

Cavallari

Lamers

Scheurwater

et al. 2013

Antimicrob Agents Chemother

103

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Pseudomonas aeruginosa is a leading cause of hospital-acquired infections and is resistant to many antibiotics. Among its primary mechanisms of resistance is expression of a chromosomally encoded AmpC ␤-lactamase that inactivates ␤-lactams. The mechanisms leading to AmpC expression in P. aeruginosa remain incompletely understood but are intricately linked to cell wall metabolism. To better understand the roles of peptidoglycan-active enzymes in AmpC expression-and consequent ␤-lactam resistance-a phenotypic screen of P. aeruginosa mutants lacking such enzymes was performed. Mutants lacking one of four lytic transglycosylases (LTs) or the nonessential penicillin-binding protein PBP4 (dacB) had altered ␤-lactam resistance. mltF and slt mutants with reduced ␤-lactam resistance were designated WIMPs (wall-impaired mutant phenotypes), while highly resistant dacB, sltB1, and mltB mutants were designated HARMs (high-level AmpC resistant mutants). Double mutants lacking dacB and sltB1 had extreme piperacillin resistance (>256 g/ml) compared to either of the single knockouts (64 g/ml for a dacB mutant and 12 g/ml for an sltB1 mutant). Inactivation of ampC reverted these mutants to wild-type susceptibility, confirming that AmpC expression underlies resistance. dacB mutants had constitutively elevated AmpC expression, but the LT mutants had wild-type levels of AmpC in the absence of antibiotic exposure. These data suggest that there are at least two different pathways leading to AmpC expression in P. aeruginosa and that their simultaneous activation leads to extreme ␤-lactam resistance.

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Section: Resultsmentioning

confidence: 99%

Section: Figmentioning

confidence: 99%

Section: Figmentioning

confidence: 99%

Section: Figmentioning

confidence: 99%

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Changes to Its Peptidoglycan-Remodeling Enzyme Repertoire Modulate β-Lactam Resistance in Pseudomonas aeruginosa

Cavallari

Lamers

Scheurwater

et al. 2013

Antimicrob Agents Chemother

103

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“…The catalytic domain is folded as in SltB1 from P. aeruginosa 13 and Slt35 from E. coli 8,14 ( Figure S4). However, the unique modular arrangement found in SltB3 provides a 27 Å-long active site that narrows (to 14.5 Å) at its exit near the proposed catalytic glutamate ( Figure 3).…”

mentioning

confidence: 99%

Turnover of Bacterial Cell Wall by SltB3, a Multidomain Lytic Transglycosylase of Pseudomonas aeruginosa

Lee¹,

Dominguez-Gil²,

Hesek³

et al. 2016

ACS Chem. Biol.

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A family of 11 lytic transglycosylases in Pseudomonas aeruginosa, an opportunistic human pathogen, turn over the polymeric bacterial cell wall in the course of its recycling, repair and maturation. The functions of these enzymes are not fully understood. We disclose herein that SltB3 of P. aeruginosa is an exolytic lytic transglycosylase. We characterize its reaction and its products by the use of peptidoglycan-based molecules. The enzyme recognizes a minimum of four sugars in its substrate, but can process a substrate comprised of a peptidoglycan of 20 sugars. The ultimate product of the reaction is N-acetylglucosamine-1,6-anhydro-N-acetylmuramic acid. The X-ray structure of this enzyme is reported for the first time. The enzyme is comprised of four domains, arranged within an annular conformation. The polymeric linear peptidoglycan substrate threads through the opening of the annulus, as it experiences turnover. Graphical AbstractCell wall, a crosslinked polymer that encases the entire bacterium, is critical for survival. The peptidoglycan, the major constituent of cell wall, is comprised of a linear polymer of repeating N-acetylglucosamine (NAG)-N-acetylmuramic acid (NAM) disaccharide, with a Corresponding Authors: mobashery@nd.edu and xjuan@iqfr.csic.es. 1 The first two authors contributed equally Supporting Information. The Supporting Information is available free of charge on the ACS Publications website. Tables S1-S2, Figures S1-S7 (PDF). The crystallographic coordinates are deposited in the Protein Data Bank (PDB codes 5anz, 5ao7 and 5ao8). (Figure 1). Crosslinking of the neighboring peptidoglycan strands takes place at the peptide stems. The mature cell wall is a single molecule of significant complexity, which is assembled and recycled in a dynamic manner. 1 Recycling of cell wall takes place in response to damage (due to antibiotics, for example), to construction of complex macromolecular assemblies such as the flagellum, or to processing during replication that requires changes to the cell wall. 2,3 The breakdown of the peptidoglycan for the recycling events is performed by lytic transglycosylases (LTs), enzymes that exist within the periplasmic space of Gram-negative bacteria in either soluble or membrane-bound forms. These enzymes perform important biological functions, which are the subjects of intense study. The importance of their functions is underscored by their multiplicity-Escherichia coli has eight 3-5 and Pseudomonas aeruginosa has eleven 6 -and by redundancies of the reactions that they perform. For example, six of the enzymes of E. coli could be ablated without any apparent consequence, but the organism cannot survive the loss of seven. 4 The mechanism of reaction of LT has been studied previously. 3,7,8 These enzymes bind to the peptidoglycan and degrade it either by removal of a disaccharide from the ends of the linear polymer (the exolytic reaction) or by cleavage of the polymer in the middle (the endolytic reaction). Whereas the catalytic sites of LTs bear resemblance to the...

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Macromolecular Machineries in Cell‐Wall Recycling

Batuecas

Hermoso

2019

Encyclopedia of Life Sciences

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Peptidoglycan, the major constituent of bacterial cell wall, is a huge polymeric macromolecule composed of glycan strands covalently linked by interconnecting peptide stems and critical for survival of bacteria. Peptidoglycan is constantly edited, biosynthesised and degraded, during essential bacterial processes such as division, elongation or insertion of the cellular machinery (e.g. flagella, pili) into cell wall. Degradation fragments are recovered, especially in Gram‐negative bacteria, by active transport and recycled to rebuild the wall. In some important Gram‐negative pathogens, β‐lactam resistance is directly linked to peptidoglycan recycling. The recycling process requires orchestration of different lytic enzymes and transporters from periplasm, the inner membrane, to cytoplasm. Recent structural and functional studies have provided relevant insights into enzymatic regulation, substrate specificity and activity of the lytic machineries involved in recycling with relevant implications in antibiotics resistance. Key Concepts Peptidoglycan recycling is an essential process in Gram‐negative bacteria that recovers self‐degradation products of cell wall in the cytoplasm to reutilise them to rebuild the wall. PG recycling is important in cell‐wall biosynthesis and in bacterial communication in many bacteria, and in antibiotics resistance in some pathogens from Enterobacteriaceae and Pseudomonadaceae families. PG recycling involves a large number of enzymes and transporters spanning from periplasm, inner and outer membranes, and cytoplasm. Lytic transglycosylases (LTs) are periplasmic and, typically, modular enzymes that initiate PG recycling by non‐hydrolytic degradation of glycan chains. They are classified in six families. Several LTs are found in each Gram‐negative bacterium. The Slt structure reveals how, under the effect of β‐lactam antibiotics, the cell wall is repaired by degradation of aberrant nascent PG chains and how it initiates the resistance phenotype. MltF is a modular membrane‐bound LT that experiences allosteric activation triggered by muropeptides. PG amidases are present in periplasm and cytoplasm. Activity in periplasm is regulated by cellular location, protein–protein interactions and oligomeric state. An activation mechanism has been discovered for cytosolic enzymes. Anhydro‐muropeptides are the key molecules to activate the AmpR to induce AmpC β‐lactamase in response to β‐lactam challenge. Regulation (in space and time) of the catalytic processes as well as of interaction between different partners is essential in PG recycling enzymes.

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Calcium-Dependent Complex Formation Between PBP2 and Lytic Transglycosylase SltB1 ofPseudomonas aeruginosa

Cited by 26 publications

References 29 publications

Changes to Its Peptidoglycan-Remodeling Enzyme Repertoire Modulate β-Lactam Resistance in Pseudomonas aeruginosa

Changes to Its Peptidoglycan-Remodeling Enzyme Repertoire Modulate β-Lactam Resistance in Pseudomonas aeruginosa

Turnover of Bacterial Cell Wall by SltB3, a Multidomain Lytic Transglycosylase of Pseudomonas aeruginosa

Macromolecular Machineries in Cell‐Wall Recycling

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