Regulated proteolysis of a cross-link–specific peptidoglycan hydrolase contributes to bacterial morphogenesis

Singh, Santosh K.; Parveen, Sadiya; SaiSree, L.; Reddy, Manjula

doi:10.1073/pnas.1507760112

Cited by 126 publications

(214 citation statements)

References 43 publications

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“…While this might imply a high degree of functional redundancy, it does not exclude the possibility that some or all of these enzymes may have unique functions under other more specific conditions. A recent study (Singh et al 2012) confirmed muramyl endopeptidase activity for three additional E. coli proteins; Spr, YebA and YdhO, renamed in E. coli to MepS, MepM and MepH (Singh et al 2015). More specifically the study presented data implying that Spr or YebA might represent endopeptidases with less redundant functions in that it was feasible to construct an Δ sprΔyebA mutant only in an E. coli strain genetically complemented with spr (mepS) (Singh et al 2012).…”

Section: Introductionmentioning

confidence: 74%

“…In this, we selected and genetically confirmed that Δ spr-mediated vancomycin sensitization, even the more pronounced sensitization of the Δ sprΔyebA mutant, required a tsp-proficient genetic background. The gene tsp itself codes for periplasmic endopeptidase implicated in processing of Spr and PBP3 (Singh et al 2015;H Hara et al 1991;H. Hara et al 1989).…”

Section: Discussionmentioning

confidence: 99%

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Muramyl endopeptidase Spr contributes to intrinsic vancomycin resistance inSalmonella entericaserovar Typhimurium

Vestoe

Huseby

Wang

et al. 2018

Preprint

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The impermeability barrier provided by the outer membrane of enteric bacteria, a feature lacking in Gram-positive bacteria, plays a major role in maintaining resistance to numerous antimicrobial compounds and antibiotics. Here we demonstrate that mutational inactivation of spr, coding for a muramyl endopeptidase, significantly sensitizes Salmonella enterica serovar Typhimurium to vancomycin without any accompanying apparent growth defect or outer membrane destabilization. A similar phenotype was not achieved by deleting the mepA, pbpG, nlpC, yebA or yhdO genes coding for functional homologues to Spr. The spr mutant revealed increased autolytic behavior in response to not only vancomycin, but also to penicillin G, an antibiotic for which the mutant displayed a wild-type MIC. A screen for suppressor mutations of the spr mutant phenotype revealed that deletion of tsp (prc), encoding a periplasmic carboxypeptidase involved in processing Spr and PBP3, restored intrinsic resistance to vancomycin and reversed the autolytic phenotype of an spr mutant. Our data suggest that Spr contributes to intrinsic antibiotic resistance in S. Typhimurium without directly affecting the outer membrane permeability barrier. Furthermore, our data suggests that compounds that target specific cell wall endopeptidases might have the potential to expand the activity spectrum of traditional Gram-positive antibiotics.

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Section: Introductionmentioning

confidence: 74%

Section: Discussionmentioning

confidence: 99%

Muramyl endopeptidase Spr contributes to intrinsic vancomycin resistance inSalmonella entericaserovar Typhimurium

Vestoe

Huseby

Wang

et al. 2018

Preprint

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“…The EMBO Journal Proteolytic activation of PG hydrolase Helene Botella et al (Singh et al, 2015). However, unlike the action of MarP on RipA, PrC controls the level of MepS in E. coli by degrading it when it is no longer needed, such as in stationary phase.…”

Section: Discussionmentioning

confidence: 99%

Mycobacterium tuberculosis protease MarP activates a peptidoglycan hydrolase during acid stress

et al. 2017

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(Mtb) can persist in the human host in a latent state for decades, in part because it has the ability to withstand numerous stresses imposed by host immunity. Prior studies have established the essentiality of the periplasmic protease MarP for Mtb to survive in acidified phagosomes and establish and maintain infection in mice. However, the proteolytic substrates of MarP that mediate these phenotypes were unknown. Here, we used biochemical methods coupled with supravital chemical probes that facilitate imaging of nascent peptidoglycan to demonstrate that during acid stress MarP cleaves the peptidoglycan hydrolase RipA, a process required for RipA's activation. Failure of RipA processing in MarP-deficient cells leads to cell elongation and chain formation, a hallmark of progeny cell separation arrest. Our results suggest that sustaining peptidoglycan hydrolysis, a process required for cell elongation, separation of progeny cells, and cell wall homeostasis in growing cells, may also be essential for Mtb's survival in acidic conditions.

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“…Much less is known about EP regulation in Gram-negative bacteria, where EPs appear to be kept in check as part of multiprotein complexes (40, 41), at the transcriptional level (42, 43) or via proteolytic degradation (44, 45). Proteolytic turnover appears to be the major mode of regulation of growth-promoting EPs (44, 45); however, phenotypes associated with the accumulation of EPs have surprisingly mild phenotypes under normal growth conditions, suggesting an additional, unexplored layer of regulation.…”

Section: Introductionmentioning

confidence: 99%

“…The Gram-positive bacterium Bacillus subtilis (and likely other Gram-positive species as well) regulates EP activity via interactions with FtsEX and the PG synthesis elongation machinery (35–37) and additionally encodes a post-translational negative regulator, IseA (YoeB) (38), which presumably serves to tone down EP activity during cell wall stress conditions (38, 39). Much less is known about EP regulation in Gram-negative bacteria, where EPs appear to be kept in check as part of multiprotein complexes (40, 41), at the transcriptional level (42, 43) or via proteolytic degradation (44, 45). Proteolytic turnover appears to be the major mode of regulation of growth-promoting EPs (44, 45); however, phenotypes associated with the accumulation of EPs have surprisingly mild phenotypes under normal growth conditions, suggesting an additional, unexplored layer of regulation.…”

Section: Introductionmentioning

confidence: 99%

Structural basis of peptidoglycan endopeptidase regulation

Shin

Sulpizio

Kelley

et al. 2019

Preprint

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AbstractMost bacteria surround themselves with a cell wall, a strong meshwork consisting primarily of the polymerized aminosugar peptidoglycan (PG). PG is essential for structural maintenance of bacterial cells, and thus for viability. PG is also constantly synthesized and turned over, the latter process is mediated by PG cleavage enzymes, for example the endopeptidases (EPs). EPs themselves are essential for growth, but also promote lethal cell wall degradation after exposure to antibiotics that inhibit PG synthases (e.g., β-lactams). Thus, EPs are attractive targets for novel antibiotics and their adjuvants. However, we have a poor understanding of how these enzymes are regulated in vivo, depriving us of novel pathways for the development of such antibiotics. Here, we have solved crystal structures of the LysM/M23 family peptidase ShyA, the primary EP of the cholera pathogen Vibrio cholerae. Our data suggest that ShyA assumes two drastically different conformations; a more open form that allows for substrate binding, and a closed form, which we predicted to be catalytically inactive. Mutations expected to promote the open conformation caused enhanced activity in vitro and in vivo, and these results were recapitulated in EPs from the divergent pathogens Neisseria gonorrheae and Escherichia coli. Our results suggest that LysM/M23 EPs are regulated via release of the inhibitory Domain1 from the M23 active site, likely through conformational re-arrangement in vivo.SignificanceBacteria digest their cell wall following exposure to antibiotics like penicillin. The endopeptidases (EPs) are among the proteins that catalyze cell wall digestion processes after antibiotic exposure, but we do not understand how these enzymes are regulated during normal growth. Herein, we present the structure of the major EP from the diarrheal pathogen Vibrio cholerae. Surprisingly, we find that EPs from this and other pathogens appear to be produced as a largely inactive precursor that undergoes a conformational shift exposing the active site to engage in cell wall digestion. These results enhance our understanding of how EPs are regulated and could open the door for the development of novel antibiotics that overactivate cell wall digestion processes.

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Regulated proteolysis of a cross-link–specific peptidoglycan hydrolase contributes to bacterial morphogenesis

Cited by 126 publications

References 43 publications

Muramyl endopeptidase Spr contributes to intrinsic vancomycin resistance inSalmonella entericaserovar Typhimurium

Muramyl endopeptidase Spr contributes to intrinsic vancomycin resistance inSalmonella entericaserovar Typhimurium

Mycobacterium tuberculosis protease MarP activates a peptidoglycan hydrolase during acid stress

Structural basis of peptidoglycan endopeptidase regulation

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