Dataset Information

Lytic transglycosylase MltG cleaves in nascent peptidoglycan and produces short glycan strands.

ABSTRACT: Bacteria encase their cytoplasmic membrane with peptidoglycan (PG) to maintain the shape of the cell and protect it from bursting. The enlargement of the PG layer is facilitated by the coordinated activities of PG synthesising and -cleaving enzymes. In Escherichia coli, the cytoplasmic membrane-bound lytic transglycosylase MltG associates with PG synthases and was suggested to terminate the polymerisation of PG glycan strands. Using pull-down and surface plasmon resonance, we detected interactions between MltG from Bacillus subtilis and two PG synthases; the class A PBP1 and the class B PBP2B. Using in vitro PG synthesis assays with radio-labelled or fluorophore-labelled B. subtilis-type and/or E. coli-type lipid II, we showed that both, BsMltG and EcMltG, are lytic tranglycosylases and that their activity is higher during ongoing glycan strand polymerisation. MltG competed with the transpeptidase activity of class A PBPs, but had no effect on their glycosyltransferase activity, and produced glycan strands with a length of 7 disaccharide units from cleavage in the nascent strands. We hypothesize that MltG cleaves the nascent strands to produce short glycan strands that are used in the cell for a yet unknown process.

SUBMITTER: Sassine J

PROVIDER: S-EPMC8135044 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature

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Lytic transglycosylase MltG cleaves in nascent peptidoglycan and produces short glycan strands.

Sassine Jad J Pazos Manuel M Breukink Eefjan E Vollmer Waldemar W

Cell surface (Amsterdam, Netherlands) 20210501

Bacteria encase their cytoplasmic membrane with peptidoglycan (PG) to maintain the shape of the cell and protect it from bursting. The enlargement of the PG layer is facilitated by the coordinated activities of PG synthesising and -cleaving enzymes. In <i>Escherichia coli</i>, the cytoplasmic membrane-bound lytic transglycosylase MltG associates with PG synthases and was suggested to terminate the polymerisation of PG glycan strands. Using pull-down and surface plasmon resonance, we detected int ...[more]

PMID: 34036206

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Project description:The ellipsoid shape of Streptococcus pneumoniae is determined by the synchronized actions of the elongasome and the divisome, which have the task of creating a protective layer of peptidoglycan (PG) enveloping the cell membrane. The elongasome is necessary for expanding PG in the longitudinal direction whereas the divisome synthesizes the PG that divides one cell into two. Although there is still little knowledge about how these two modes of PG synthesis are coordinated, it was recently discovered that two RNA-binding proteins called EloR and KhpA are part of a novel regulatory pathway controlling elongation in S. pneumoniae EloR and KhpA form a complex that work closely with the Ser/Thr kinase StkP to regulate cell elongation. Here, we have further explored how this regulation occur. EloR/KhpA is found at midcell, a localization fully dependent on EloR. Using a bacterial two-hybrid assay we probed EloR against several elongasome proteins and found an interaction with the lytic transglycosylase homolog MltG. By using EloR as bait in immunoprecipitation assays, MltG was pulled down confirming that they are part of the same protein complex. Fluorescent microscopy demonstrated that the Jag domain of EloR is essential for EloR's midcell localization and its interaction with MltG. Since MltG is found at midcell independent of EloR, our results suggest that MltG is responsible for recruitment of the EloR/KhpA complex to the division zone to regulate cell elongation.Importance Bacterial cell division has been a successful target for antimicrobial agents for decades. How different pathogens regulate cell division is, however, poorly understood. To fully exploit the potential for future antibiotics targeting cell division, we need to understand the details of how the bacteria regulate and construct cell wall during this process. Here we have revealed that the newly identified EloR/KhpA complex, regulating cell elongation in S. pneumoniae, forms a complex with the essential peptidoglycan transglycosylase MltG at midcell. EloR, KhpA and MltG are conserved among many bacterial species and the EloR/KhpA/MltG regulatory pathway is most likely a common mechanism employed by many Gram-positive bacteria to coordinate cell elongation and septation.

Project description:Chlamydiales species are obligate intracellular bacteria lacking a classical peptidoglycan sacculus but relying on peptidoglycan synthesis for cytokinesis. While septal peptidoglycan biosynthesis seems to be regulated by MreB actin and its membrane anchor RodZ rather than FtsZ tubulin in Chlamydiales, the mechanism of peptidoglycan remodeling is poorly understood. An amidase conserved in Chlamydiales is able to cleave peptide stems in peptidoglycan, but it is not clear how peptidoglycan glycan strands are cleaved since no classical lytic transglycosylase is encoded in chlamydial genomes. However, a protein containing a SpoIID domain, known to possess transglycosylase activity in Bacillus subtilis, is conserved in Chlamydiales We show here that the SpoIID homologue of the Chlamydia-related pathogen Waddlia chondrophila is a septal peptidoglycan-binding protein. Moreover, we demonstrate that SpoIID acts as a lytic transglycosylase on peptidoglycan and as a muramidase on denuded glycan strands in vitro As SpoIID-like proteins are widespread in nonsporulating bacteria, SpoIID might commonly be a septal peptidoglycan remodeling protein in bacteria, including obligate intracellular pathogens, and thus might represent a promising drug target.IMPORTANCEChlamydiales species are obligate intracellular bacteria and important human pathogens that have a minimal division machinery lacking the proteins that are essential for bacterial division in other species, such as FtsZ. Chlamydial division requires synthesis of peptidoglycan, which forms a ring at the division septum and is rapidly turned over. However, little is known of peptidoglycan degradation, because many peptidoglycan-degrading enzymes are not encoded by chlamydial genomes. Here we show that an homologue of SpoIID, a peptidoglycan-degrading enzyme involved in sporulation of bacteria such as Bacillus subtilis, is expressed in Chlamydiales, localizes at the division septum, and degrades peptidoglycan in vitro, indicating that SpoIID is not only involved in sporulation but also likely implicated in division of some bacteria.

Dataset Information

Lytic transglycosylase MltG cleaves in nascent peptidoglycan and produces short glycan strands.

Publications

Lytic transglycosylase MltG cleaves in nascent peptidoglycan and produces short glycan strands.

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