β-Barrels covalently link peptidoglycan and the outer membrane in the α-proteobacterium Brucella abortus

Godessart, Pierre; Lannoy, Adélie; Dieu, Marc; Verren, Sander E Van der; Soumillion, Patrice; Collet, Jean-François; Remaut, Han; Renard, Patricia; Bolle, Xavier De

doi:10.1038/s41564-020-00799-3

“…Some of the DpaA homologues might function as LD-carboxypeptidase, consistent with their classification in the MEROPS peptidase database (43) within the C82.A01 peptidase subfamily along with the LD-carboxypeptidases Csd6 from Helicobacter pylori and Pgp2 from Campylobacter jejuni (44,45). Alternatively, or in addition, DpaA could detach different substrates from PG, for example OMPs in Alphaand Gammaproteobacteria (24,25). Indeed, a search within the genomes of Alphaand Gammaproteobacteria with PG-attached OMPs identified genes encoding DpaA-like proteins in all of them (Fig.…”

Section: Resultsmentioning

confidence: 52%

“…While another PGattached lipoprotein has been described for Pseudomonas spp., the covalent attachment of PG to OM-localized proteins has not been studied in depth for PG of other diderm bacteria (23). This changed recently when two publications reported the covalent attachment of certain OM b-barrel proteins (OMPs) in a wide range of Alphaproteobacteria (24,25). Additionally to OMPs, one of the studies identified the OM-anchored lipoprotein LimB attached to PG in Coxiella burnetii (25).…”

mentioning

confidence: 99%

DpaA Detaches Braun’s Lipoprotein from Peptidoglycan

Winkle

¹

,

Hernández-Rocamora

²

,

Pullela

³

et al. 2021

mBio

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Gram-negative bacteria have a unique cell envelope with a lipopolysaccharide-containing outer membrane that is tightly connected to a thin layer of peptidoglycan. The tight connection between the outer membrane and peptidoglycan is needed to maintain the outer membrane as an impermeable barrier for many toxic molecules and antibiotics. Enterobacteriaceae such as Escherichia coli covalently attach the abundant outer membrane-anchored lipoprotein Lpp (Braun’s lipoprotein) to tripeptides in peptidoglycan, mediated by the transpeptidases LdtA, LdtB, and LdtC. LdtD and LdtE are members of the same family of ld-transpeptidases but they catalyze a different reaction, the formation of 3-3 cross-links in the peptidoglycan. The function of the sixth homologue in E. coli, LdtF, remains unclear, although it has been shown to become essential in cells with inhibited lipopolysaccharide export to the outer membrane. We now show that LdtF hydrolyzes the Lpp-peptidoglycan linkage, detaching Lpp from peptidoglycan, and have renamed LdtF to peptidoglycan meso-diaminopimelic acid protein amidase A (DpaA). We show that the detachment of Lpp from peptidoglycan is beneficial for the cell under certain stress conditions and that the deletion of dpaA allows frequent transposon inactivation in the lapB (yciM) gene, whose product downregulates lipopolysaccharide biosynthesis. DpaA-like proteins have characteristic sequence motifs and are present in many Gram-negative bacteria, of which some have no Lpp, raising the possibility that DpaA has other substrates in these species. Overall, our data show that the Lpp-peptidoglycan linkage in E. coli is more dynamic than previously appreciated. IMPORTANCE Gram-negative bacteria have a complex cell envelope with two membranes and a periplasm containing the peptidoglycan layer. The outer membrane is firmly connected to the peptidoglycan by highly abundant proteins. The outer membrane-anchored Braun’s lipoprotein (Lpp) is the most abundant protein in E. coli, and about one-third of the Lpp molecules become covalently attached to tripeptides in peptidoglycan. The attachment of Lpp to peptidoglycan stabilizes the cell envelope and is crucial for the outer membrane to function as a permeability barrier for a range of toxic molecules and antibiotics. So far, the attachment of Lpp to peptidoglycan has been considered to be irreversible. We have now identified an amidase, DpaA, which is capable of detaching Lpp from peptidoglycan, and we show that the detachment of Lpp is important under certain stress conditions. DpaA-like proteins are present in many Gram-negative bacteria and may have different substrates in these species.

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“…Some of the DpaA homologues might function as LD-carboxypeptidase, consistent with their classification in the MEROPS peptidase database (39) within the C82.A01 peptidase subfamily along with the LD-carboxypeptidases Csd6 from Helicobacter pylori and Pgp2 from Campylobacter jejuni (40, 41). Alternatively, or in addition, DpaA could detach different substrates from PG, for example OMPs in α- and γ-proteobacteria (24, 25). Indeed, a search within the genomes of α- and γ-proteobacteria with PG-attached OMPs identified genes encoding DpaA-like proteins in all of them ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…However, at present we cannot exclude the possibility that some bacteria employ DpaA-like enzymes to remove non-canonical D-amino acids, which are toxic when incorporated at a high amount (47). The recent discovery of PG-attached OMPs in certain α-proteobacteria suggests that the DpaA-like enzymes in these species function to detach OMPs from PG (24, 25). In support of this hypothesis, the attachment of OMPs to PG occurs via N-terminal glycine or alanine, and the attachment of OM-lipoprotein LimB via an internal lysine residue, producing the very same amide bonds as in TetraGly4 or in PG-bound Lpp in E. coli .…”

Section: Discussionmentioning

confidence: 99%

“…While another PG-attached lipoprotein has been described in Pseudomonas spp., the covalent attachment of PG to OM localized proteins has not been studied in depth in other diderm bacteria PG (23). This changed recently when two publications reported the covalent attachment of certain OM β-barrel proteins (OMPs) in a wide range of α-proteobacteria (24, 25). Additionally to OMPs, one of the studies identified the OM-anchored lipoprotein LimB attached to PG in Coxiella burnetti (25).…”

Section: Introductionmentioning

confidence: 99%

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DpaA detaches Braun’s lipoprotein from peptidoglycan

Winkle

¹

,

Hernández-Rocamora

²

,

Pullela

³

et al. 2021

Preprint

View full text Add to dashboard Cite

Gram-negative bacteria have a unique cell envelope with a lipopolysaccharide-containing outer membrane that is tightly connected to a thin layer of peptidoglycan. The tight connection between the outer membrane and peptidoglycan is needed to maintain the outer membrane as an impermeable barrier for many toxic molecules and antibiotics. Enterobacteriaceae such as Escherichia coli covalently attach the abundant outer membrane-anchored lipoprotein Lpp (Braun's lipoprotein) to tripeptides in peptidoglycan, mediated by the transpeptidases LdtA, LdtB and LdtC. LdtD and LdtE are members of the same family of LD-transpeptidases but they catalyse a different reaction, the formation of 3-3 cross-links in the peptidoglycan. The function of the sixth homologue in E. coli, LdtF remains unclear, although it has been shown to become essential in cells with inhibited LPS export to the outer membrane. We now show that LdtF hydrolyses the Lpp-peptidoglycan linkage, detaching Lpp from peptidoglycan, and have renamed LdtF to peptidoglycan meso-diaminopimelic acid protein amidase A (DpaA). We show that the detachment of Lpp from peptidoglycan is beneficial for the cell under certain stress conditions and that the deletion of dpaA allows frequent transposon inactivation in the lapB (yciM) gene, whose product down-regulates lipopolysaccharide biosynthesis. DpaA-like proteins have characteristic sequence motifs and are present in many Gram-negative bacteria of which some have no Lpp, raising the possibility that DpaA has other substrates in these species. Overall, our data show that the Lpp-peptidoglycan linkage in E. coli is more dynamic than previously appreciated.

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