Regulation of peptidoglycan hydrolases: localization, abundance, and activity

Brogan, Anna P.; Rudner, David Z.

doi:10.1016/j.mib.2023.102279

Cited by 16 publications

(14 citation statements)

References 73 publications

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“…These proteins ensure the proper packaging and condensation of DNA, as well as the assembly of the highly resistant outer layers that protect the spore’s genetic material (11). On the other hand, the hydrolase-related proteins identified in spores serve a critical function during the germination process - the transition from the dormant spore state back to a metabolically active vegetative cell (52). The predominant proteins were further examined by KEGG pathway analysis which showed proteins involved in peptidoglycan (PG) biosynthesis, glycolysis, carbon metabolism and biosynthesis of secondary metabolites are shared between cells and spores albeit that the relative quantities of the proteins vary in both.…”

Section: Discussionmentioning

confidence: 99%

“…The predominant proteins were further examined by KEGG pathway analysis which showed proteins involved in peptidoglycan (PG) biosynthesis, glycolysis, carbon metabolism and biosynthesis of secondary metabolites are shared between cells and spores albeit that the relative quantities of the proteins vary in both. Both the spores and cells of B. subtilis are encased in a robust PG layer, offering structural stability and a shield against environmental adversities (52, 53). During sporulation, a layer of PG cortex is formed, crucial for spore dehydration and reinforcing spore resistance.…”

Section: Discussionmentioning

confidence: 99%

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Non-mechanical disruption ofBacillus subtilis sporesallows sensitive and deep characterization of the minimal proteome for resuming a cellular lifestyle

Huang,

de Koster,

et al. 2024

Preprint

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In response to extreme conditions, Bacillus subtilis generates highly resilient spores characterized by a unique multilayered structure. This confers resistance against various chemicals and enzymes yet adding complexity to the analysis of the spore proteome. As the first step in bottom-up proteomics, sample preparation poses a significant challenge. We assessed how an optimized protocol for sample preparation by easy extraction and digestion (SPEED) performed compared to previously established methods "One-pot" (OP) and single-pot, solid phase-enhanced sample-preparation (SP3) for the proteomic analysis of B. subtilis cell and spore samples. We found that SPEED outperformed both OP and SP3 in terms of peptides and proteins identified, moreover SPEED highly reproducibly quantified over 1000 proteins in limited input samples as low as 1 OD600 of B. subtilis cells and spores. SPEED was applied to analyze spore samples of different purity by applying sequential purification following harvesting of spores. Comparison of the differential abundance of proteins revealed clusters likely partially stemming from remaining vegetative cells in less purified spore samples. We show that ranking of absolute protein abundance in cellular and spore samples further enables us to rationally differentiate integral spore proteins from vegetative remnants. This is of importance in applications and organisms where highly homogenous spore samples are difficult to obtain. A deep proteomic analysis of spore and vegetative cell samples with the new approach led to the identification of 2447 proteins, 2273 of which were further quantified and compared between B. subtilis spores and cells. Our findings indicate that pathways related to peptidoglycan biosynthesis, glycolysis, carbon metabolism, and biosynthesis of secondary metabolites are shared between cells and spores. This corroborates and extends earlier work stressing that despite marked differences in their physiological states, spores preserve vegetative cell (core) proteins, essential for revival under conditions conducive to growth.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Non-mechanical disruption ofBacillus subtilis sporesallows sensitive and deep characterization of the minimal proteome for resuming a cellular lifestyle

Huang,

de Koster,

et al. 2024

Preprint

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“…These results are consistent with key peptidoglycan hydrolases in other bacterial species, such as AmiA/B in Escherichia coli 24 , CwlO/LytE in Bacillus subtilis 25,26 , PcsB in Streptococcus pneumoniae 27 and RipC in Mycobacterium tuberculosis 28 . Although the mechanisms of regulation may differ amongst peptidoglycan hydrolases and remain to be determined for SagA, the sagA promoter contains sequence motifs that may be regulated by WalRK two-component systems 29 and the SagA protein contains a N-terminal coil-coil domain that may interact with E. faecium divisome components, which warrants further investigation in the future. Notably, the deletion of sagA in E. faecium Com15 also renders this strain more susceptible to cell wall-acting antibiotics ( Extended Data Fig.…”

Section: Main Textmentioning

confidence: 99%

Secreted antigen A peptidoglycan hydrolase is essential forEnterococcus faeciumcell separation and priming of immune checkpoint inhibitor therapy

Klupt,

Fam,

Zhang

et al. 2023

Preprint

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Introductory paragraphEnterococcus faeciumis a microbiota species in humans that can modulate host immunity1, but has also acquired antibiotic resistance and is a major cause of hospital-associated infections2. Notably, diverse strains ofE. faeciumproduce SagA, a highly conserved peptidoglycan hydrolase that is sufficient to promote intestinal immunity3–5and immune checkpoint inhibitor antitumor activity6. However, the essential functions of SagA inE. faeciumwere unknown. Here we report that deletion ofsagAimpairedE. faeciumgrowth and resulted in bulged and clustered enterococci due to defective peptidoglycan cleavage and cell separation. Moreover, ΔsagAshowed increased antibiotic sensitivity, yielded lower levels of active muropeptides, displayed reduced activation of the peptidoglycan pattern-recognition receptor NOD2, and failed to promote cancer immunotherapy. Importantly, plasmid-based expression of SagA, but not its catalytically-inactive mutant, restored ΔsagAgrowth, production of active muropeptides and NOD2 activation. SagA is therefore essential forE. faeciumgrowth, stress resistance and activation of host immunity.

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“…These latter two enzymatic activities are either provided by bifunctional class A penicillin binding proteins (PBPs) or by SEDS-type glycosyltransferases that cooperate with class B PBPs that are mere transpeptidases (8)(9)(10)(11). The PG network is very dynamic and also remodelled by degradative enzymes to adjust size and shape to growth, division and development (12). PG biosynthesis is fuelled either with UDP-GlcNAc generated by the GlmSMU enzymes from fructose-6-phosphate, an intermediate of glycolysis (4), or by salvage of PG precursors from environmental sources (13).…”

Section: Introductionmentioning

confidence: 99%

Cytosolic factors controlling PASTA kinase-dependent ReoM phosphorylation

Rothe,

Wamp,

Rosemeyer

et al. 2024

Preprint

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Bacteria adapt the biosynthesis of their envelopes to specific growth conditions and prevailing stress factors. Peptidoglycan (PG) is the major component of the cell wall in Gram-positive bacteria, where PASTA kinases play a central role in PG biosynthesis regulation. Despite their importance for growth, cell division and antibiotic resistance, the mechanisms of PASTA kinase activation are not fully understood. ReoM, a recently discovered cytosolic phosphoprotein, is one of the main substrates of the PASTA kinase PrkA in the Gram-positive human pathogen Listeria monocytogenes. Depending on its phosphorylation, ReoM controls proteolytic stability of MurA, the first enzyme in the PG biosynthesis pathway. The late cell division protein GpsB has been implicated in PASTA kinase signalling. Consistently, we show that L. monocytogenes prkA and gpsB mutants phenocopied each other. Analysis of in vivo ReoM phosphorylation confirmed GpsB as an activator of PrkA leading to the description of structural features in GpsB that are important for kinase activation. We further show that ReoM phosphorylation is growth-phase dependent and that this kinetic is reliant on the protein phosphatase PrpC. ReoM phosphorylation was inhibited in mutants with defects in MurA degradation, leading to the discovery that artificial MurA overexpression prevented ReoM phosphorylation. Overexpressed MurA must adopt its substrate-bound closed conformation and interact with ReoM to exert this effect, but the extracellular PASTA domains of PrkA or MurJ flippases where not required. Our results indicate that intracellular signals control ReoM phosphorylation and extend current models describing the mechanisms of PASTA kinase activation.

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Regulation of peptidoglycan hydrolases: localization, abundance, and activity

Cited by 16 publications

References 73 publications

Non-mechanical disruption ofBacillus subtilis sporesallows sensitive and deep characterization of the minimal proteome for resuming a cellular lifestyle

Non-mechanical disruption ofBacillus subtilis sporesallows sensitive and deep characterization of the minimal proteome for resuming a cellular lifestyle

Secreted antigen A peptidoglycan hydrolase is essential forEnterococcus faeciumcell separation and priming of immune checkpoint inhibitor therapy

Cytosolic factors controlling PASTA kinase-dependent ReoM phosphorylation

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