Understanding Protein Glycosylation Pathways in Bacteria

Li, Hong; Debowski, Aleksandra W.; Liao, Tingting; Tang, Hong; Nilsson, Hans Olof; Marshall, Barry J.; Stubbs, Keith A.; Benghezal, Mohammed

doi:10.2217/fmb-2016-0166

Cited by 24 publications

(10 citation statements)

References 91 publications

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“…Several studies have demonstrated that glycosylation is one of the most important modifications used by P. gingivalis to increase its virulence. Bacterial glycosylation is related mainly to macromolecules exposed to the external environment [65] and is important for bacterial interactions with and adhesion to the host cells [66]. Among the best-characterized glycoconjugates are gingipains [67, 68], an outer membrane OMP85 protein [69], and Mfa1 fimbriae [70].…”

Section: Discussionmentioning

confidence: 99%

PgFur participates differentially in expression of virulence factors in more virulent A7436 and less virulent ATCC 33277 Porphyromonas gingivalis strains

et al. 2019

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Background Porphyromonas gingivalis is considered a keystone pathogen responsible for chronic periodontitis. Although several virulence factors produced by this bacterium are quite well characterized, very little is known about regulatory mechanisms that allow different strains of P. gingivalis to efficiently survive in the hostile environment of the oral cavity, a typical habitat characterized by low iron and heme concentrations. The aim of this study was to characterize P. gingivalis Fur homolog (PgFur) in terms of its role in production of virulence factors in more (A7436) and less (ATCC 33277) virulent strains. Results Expression of a pgfur depends on the growth phase and iron/heme concentration. To better understand the role played by the PgFur protein in P. gingivalis virulence under low- and high-iron/heme conditions, a pgfur -deficient ATCC 33277 strain (TO16) was constructed and its phenotype compared with that of a pgfur A7436-derived mutant strain (TO6). In contrast to the TO6 strain, the TO16 strain did not differ in the growth rate and hemolytic activity compared with the ATCC 33277 strain. However, both mutant strains were more sensitive to oxidative stress and they demonstrated changes in the production of lysine- (Kgp) and arginine-specific (Rgp) gingipains. In contrast to the wild-type strains, TO6 and TO16 mutant strains produced larger amounts of HmuY protein under high iron/heme conditions. We also demonstrated differences in production of glycoconjugates between the A7436 and ATCC 33277 strains and we found evidence that PgFur protein might regulate glycosylation process. Moreover, we revealed that PgFur protein plays a role in interactions with other periodontopathogens and is important for P. gingivalis infection of THP-1-derived macrophages and survival inside the cells. Deletion of the pgfur gene influences expression of many transcription factors, including two not yet characterized transcription factors from the Crp/Fnr family. We also observed lower expression of the CRISPR/Cas genes. Conclusions We show here for the first time that inactivation of the pgfur gene exerts a different influence on the phenotype of the A7436 and ATCC 33277 strains. Our findings further support the hypothesis that PgFur regulates expression of genes encoding surface virulence factors and/or genes involved in their maturation. Electronic supplementary material The online version of this article (10.1186/s12866-019-1511-x) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

PgFur participates differentially in expression of virulence factors in more virulent A7436 and less virulent ATCC 33277 Porphyromonas gingivalis strains

et al. 2019

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“…1). In most wellcharacterized systems N-glycosylation occurs in the eukaryotic endoplasmic reticulum or bacterial periplasm and requires an oligosaccharyltransferase (OST) to transfer a preassembled oligosaccharide from a lipid donor to asparagine residues in substrate proteins (43)(44)(45). In contrast, NTHi employs HMW-C, a soluble cytoplasmic protein belonging to the GT41 family of glycosyltransferases, a family which is otherwise comprised of O-GlcNAc transferases (46).…”

Section: Introductionmentioning

confidence: 99%

Phase-Variable Glycosylation in Nontypeable Haemophilus influenzae

Elango

Schulz

2019

J. Proteome Res.

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Non-typeable Haemophilus influenzae (NTHi) is a leading cause of respiratory tract infections worldwide and continues to be a global health burden. Adhesion and colonisation of host cells are crucial steps in bacterial pathogenesis, and in many strains of NTHi interaction with the host is mediated by the high molecular weight adhesins HMW1A and HMW2A. These adhesins are N-glycoproteins which are modified by cytoplasmic glycosyltransferases HMW1C and HMW2C. Phase variation in the number of short sequence repeats in the promoters of hmw1A and hmw2A directly affects their expression. Here, we report the presence of similar variable repeat elements in the promoters of hmw1C and hmw2C in diverse NTHi isolates. In an ex vivo assay, we systematically altered substrate and glycosyltransferase expression and showed that both of these factors affected the site-specific efficiency of glycosylation on HMW-A. Glycosylation occupancy was incomplete at many sites, variable between sites, and generally lower close to the C-terminus of HMW-A. We investigated the causes of this variability. As HMW-C glycosylates HMW-A in the cytoplasm, we tested how secretion affected glycosylation on HMW-A and showed that retaining HMW-A in the cytoplasm indeed increased glycosylation occupancy across the full length of the protein. Sitedirected mutagenesis showed that HMW-C had no inherent preference for glycosylating asparagines in NxS or NxT sequons. This work provides key insights into factors contributing to the heterogenous modifications of NTHi HMW-A adhesins, expands knowledge of NTHi population diversity and pathogenic capability, and is relevant to vaccine design for NTHi and related pathogens..

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“…For a long time, protein glycosylation was deemed to be restricted to eukaryotes. So far, it has been clearly established that bacteria, including pathogens possess both O-and N-linked glycosylation pathways that display many commonalities with their eukaryotic counterparts, as well as some unexpected variations (2,3). N-linked protein glycosylation commonly occurs at the side chain amide of an Asn residue in the sequences Asn-X-Ser/Thr of the targeted proteins, where X can be any amino acid except proline.…”

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confidence: 99%

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

Liu

et al. 2019

J Bacteriol

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A bacterial inverting glycosyltransferase EarP transfers rhamnose from dTDP-β-l-rhamnose (TDP-Rha) to Arg32 of translation elongation factor P (EF-P) to activate its function. We report here the structural and biochemical characterization of Pseudomonas aeruginosa EarP. In contrast to recently reported Neisseria meningitidis EarP, P. aeruginosa EarP exhibits differential conformational changes upon TDP-Rha and EF-P binding. Sugar donor binding enhances acceptor binding to EarP, as revealed by structural comparison between the apo-, TDP-Rha-, and TDP/EF-P-bound forms and isothermal titration calorimetry experiments. In vitro EF-P rhamnosylation combined with active-site geometry indicates that Asp16 corresponding to Asp20 of N. meningitidis EarP is the catalytic base, whereas Glu272 is another putative catalytic residue. Our study should provide the basis for EarP-targeted inhibitor design against infections from P. aeruginosa and other clinically relevant species. IMPORTANCE Posttranslational rhamnosylation of EF-P plays a key role in Pseudomonas aeruginosa, establishing virulence and antibiotic resistance, as well as survival. The detailed structural and biochemical characterization of the EF-P-specific rhamnosyltransferase EarP from P. aeruginosa not only demonstrates that sugar donor TDP-Rha binding enhances acceptor EF-P binding to EarP but also should provide valuable information for the structure-guided development of its inhibitors against infections from P. aeruginosa and other EarP-containing pathogens.

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Understanding Protein Glycosylation Pathways in Bacteria

Cited by 24 publications

References 91 publications

PgFur participates differentially in expression of virulence factors in more virulent A7436 and less virulent ATCC 33277 Porphyromonas gingivalis strains

PgFur participates differentially in expression of virulence factors in more virulent A7436 and less virulent ATCC 33277 Porphyromonas gingivalis strains

Phase-Variable Glycosylation in Nontypeable Haemophilus influenzae

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

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