Peptidoglycan: Structure, Synthesis, and Regulation

Garde, Shambhavi; Chodisetti, Pavan Kumar; Reddy, Manjula

doi:10.1128/ecosalplus.esp-0010-2020

Cited by 65 publications

(46 citation statements)

References 350 publications

(459 reference statements)

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“…A complementary pulldown experiment. A solution of recombinant N-terminal His-tagged RlpA-Δ32 was incubated separately with the two proteome preparations, in the presence of the protein crosslinker bis(sulfosuccinimidyl)suberate (BS 3 ). Ni-NTA resin was added to this mixture to entrap RlpA-Δ32 and partner ("prey") protein(s) in the preformed covalent complexes (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…His-tagged RlpA-Δ32 (300 µg) was incubated with 3 mg of lysate (wild-type PAO1 strain, OD 600 1.0, exponential phase, 1 mL of a 3 mg mL -1 solution membrane preparation or soluble-fraction) overnight, rotating at 4 °C. Subsequently, the mixture was incubated with 0.2 mL of a 5 mM solution (in DMSO) of the crosslinker bis(sulfosuccinimidyl)suberate (BS 3 , Thermo Fisher Scientific), by rotation at room temperature for 30 min. The reaction was quenched by setting the final buffer concentration to 50 mM Tris•HCl pH 8 buffer by addition of 100 μL of 500 mM Tris•HCl pH 8 (15 min reaction, rt).…”

Section: Methodsmentioning

confidence: 99%

“…A structurally unique stem peptide-often with a pentapeptide L-Ala-γ-D-Glu-m-DAP-D-Ala-D-Ala (where DAP is diaminopimelate)-is appended to the NAM saccharide. The stem from one glycan strand is crosslinked to that of a neighboring strand [1][2][3][4][5] . This cell wall polymer encases the cytoplasmic membrane and provides structural integrity to the bacterium.…”

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confidence: 99%

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In vitro studies of the protein-interaction network of cell-wall lytic transglycosylase RlpA of Pseudomonas aeruginosa

et al. 2022

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The protein networks of cell-wall-biosynthesis assemblies are largely unknown. A key class of enzymes in these assemblies is the lytic transglycosylases (LTs), of which eleven exist in P. aeruginosa. We have undertaken a pulldown strategy in conjunction with mass-spectrometry-based proteomics to identify the putative binding partners for the eleven LTs of P. aeruginosa. A total of 71 putative binding partners were identified for the eleven LTs. A systematic assessment of the binding partners of the rare lipoprotein A (RlpA), one of the pseudomonal LTs, was made. This 37-kDa lipoprotein is involved in bacterial daughter-cell separation by an unknown process. RlpA participates in both the multi-protein and multi-enzyme divisome and elongasome assemblies. We reveal an extensive protein-interaction network for RlpA involving at least 19 proteins. Their kinetic parameters for interaction with RlpA were assessed by microscale thermophoresis, surface-plasmon resonance, and isothermal-titration calorimetry. Notable RlpA binding partners include PBP1b, PBP4, and SltB1. Elucidation of the protein-interaction networks for each of the LTs, and specifically for RlpA, opens opportunities for the study of their roles in the complex protein assemblies intimately involved with the cell wall as a structural edifice critical for bacterial survival.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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confidence: 99%

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In vitro studies of the protein-interaction network of cell-wall lytic transglycosylase RlpA of Pseudomonas aeruginosa

et al. 2022

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“…Most bacteria build a peptidoglycan (PG) cell wall outside their cytoplasmic membrane. The rigid PG structure, or sacculus, surrounds the membrane and protects these unicellular microorganisms from osmotic lysis that would otherwise be caused by the high osmotic pressure in their cytoplasm. , Indeed, the PG cell wall is so strong that it can be isolated from other cellular components by subjecting cells to extensive boiling in the presence of detergents . The PG cell wall can also serve as a structure onto which other envelope components such as proteins and polysaccharides can be attached.…”

Section: Introductionmentioning

confidence: 99%

The Bacterial Cell Wall: From Lipid II Flipping to Polymerization

et al. 2022

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The peptidoglycan (PG) cell wall is an extra-cytoplasmic glycopeptide polymeric structure that protects bacteria from osmotic lysis and determines cellular shape. Since the cell wall surrounds the cytoplasmic membrane, bacteria must add new material to the PG matrix during cell elongation and division. The lipid-linked precursor for PG biogenesis, Lipid II, is synthesized in the inner leaflet of the cytoplasmic membrane and is subsequently translocated across the bilayer so that the PG building block can be polymerized and cross-linked by complex multiprotein machines. This review focuses on major discoveries that have significantly changed our understanding of PG biogenesis in the past decade. In particular, we highlight progress made toward understanding the translocation of Lipid II across the cytoplasmic membrane by the MurJ flippase, as well as the recent discovery of a novel class of PG polymerases, the SEDS (shape, elongation, division, and sporulation) glycosyltransferases RodA and FtsW. Since PG biogenesis is an effective target of antibiotics, these recent developments may lead to the discovery of much-needed new classes of antibiotics to fight bacterial resistance.

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“…The monomeric lipid-II moieties are flipped across the IM to the periplasmic space, and subsequently polymerized to synthesize PG either for sidewall synthesis (during cell elongation) or septal wall synthesis at the mid-cell (during cell division). Two distinct multiprotein complexes, namely, elongasome and divisome, execute the side and septal wall syntheses, respectively (reviewed in Garde et al, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

A LytM-Domain Factor, ActS, Functions in Two Distinctive Peptidoglycan Hydrolytic Pathways in E. coli

Chodisetti

Bahadur²,

Amrutha³

et al. 2022

Front. Microbiol.

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Bacterial cell wall contains peptidoglycan (PG) to protect the cells from turgor and environmental stress. PG consists of polymeric glycans cross-linked with each other by short peptide chains and forms an elastic mesh-like sacculus around the cytoplasmic membrane. Bacteria encode a plethora of PG hydrolytic enzymes of diverse specificity playing crucial roles in growth, division, or turnover of PG. In Escherichia coli, the cross-link-specific endopeptidases, MepS, -M, and -H, facilitate the enlargement of PG sacculus during cell elongation, whereas LytM-domain factors, EnvC and NlpD activate the division-specific amidases, AmiA, -B, and -C to facilitate the cell separation. In a screen to isolate additional factors involved in PG enlargement, we identified actS (encoding a LytM paralog, formerly ygeR) as its overexpression compensated the loss of elongation-specific endopeptidase, MepS. The overexpression of ActS resulted in the generation of partly denuded glycan strands in PG sacculi, indicating that ActS is either an amidase or an activator of amidase(s). The detailed genetic and biochemical analyses established that ActS is not a PG hydrolase, but an activator of the division-specific amidase, AmiC. However, interestingly, the suppression of the mepS growth defects by actS is not mediated through AmiC. The domain-deletion experiments confirmed the requirement of the N-terminal LysM domain of ActS for the activation of AmiC, but not for the alleviation of growth defects in mepS mutants, indicating that ActS performs two distinctive PG metabolic functions. Altogether our results suggest that in addition to activating the division-specific amidase, AmiC, ActS modulates yet another pathway that remains to be identified.

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Peptidoglycan: Structure, Synthesis, and Regulation

Cited by 65 publications

References 350 publications

In vitro studies of the protein-interaction network of cell-wall lytic transglycosylase RlpA of Pseudomonas aeruginosa

In vitro studies of the protein-interaction network of cell-wall lytic transglycosylase RlpA of Pseudomonas aeruginosa

The Bacterial Cell Wall: From Lipid II Flipping to Polymerization

A LytM-Domain Factor, ActS, Functions in Two Distinctive Peptidoglycan Hydrolytic Pathways in E. coli

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