A Second β-Hexosaminidase Encoded in the Streptococcus pneumoniae Genome Provides an Expanded Biochemical Ability to Degrade Host Glycans

Robb, Melissa; Robb, C.S.; Higgins, M.A.; Hobbs, Joanne K.; Paton, James C.; Boraston, A.B.

doi:10.1074/jbc.m115.688630

Cited by 22 publications

(34 citation statements)

References 53 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…pneumoniae -host interaction. SP_2141 and SP_2144 have been functionally studied and shown to be glycoside hydrolases (GH) with exo -β- N -acetylhexosaminidase and α-(1,6)-mannosidase activity, respectively [21, 28]. GHs are classified into >133 amino acid sequence-based families and SP_2144 falls into GH family 125, thus it is referred to as SpGH125, whereas SP_2141 (SpGH20C) falls into GH family 20.…”

Section: Resultsmentioning

confidence: 99%

“…pneumoniae whose protein products are putatively associated with N-glycan processing. The CPL consists of a characterized α-(1,6)-mannosidase (SpGH125) [21], two putative α-mannosidases (SpGH38 and SpGH92), a predicted α-fucosidase (GH29), a putative sugar kinase (ROK) and a known exo -β-hexosaminidase (SpGH20C) [28]. Other conserved proteins predicted to be associated with N-glycan processing include a characterized endo-β-N-acetylglucosaminidase (SpGH85 or EndoD) that is active on the core of N-linked glycans [24] and an ABC transporter (ABC NG ) that consists of a predicted solute binding protein (NgtS) and two putative permeases (NgtP1 and NgtP2).…”

Section: Resultsmentioning

confidence: 99%

“…Given the predicted and known activities of these enzymes, respectively (Table 1), they may be involved in degrading the carbohydrate motifs that occasionally cap the arms of complex N-glycans, such as the histo-blood group antigens. Indeed, our recent work on SpGH20C indicates that this is a generalist exo-β-N-acetylhexosaminidase that may have the ability to process terminal β-linked GlcNAc or GalNAc residues in such capping motifs [28]. Alternatively, the function of CPL as a whole may not be specifically targeted at N-glycans and these additional GHs may target the linkages in O-glycans.…”

Section: Discussionmentioning

confidence: 99%

“…For western blotting, TIGR4 Sm r was grown in AGCH media containing 1% mannose, glucose or galactose at 37°C for 10 hours, pelleted and resuspended in Laemmli buffer. Samples were heated, run on a 10% SDS-PAGE gel and western blotting performed as previously described [28] using a 1/4000 dilution of rabbit antiserum raised against purified recombinant SpGH92. For FACE analysis, TIGR4 Sm r was grown in 50 mL AGCH medium with 1% glucose to an OD 600 of 0.6, then pelleted and resuspended in AGCH containing no sugar and incubated for a further 30 minutes (in an attempt to induce expression of SpGH92).…”

Section: Methodsmentioning

confidence: 99%

“…Lane 1–4: 100, 50, 10 and 1 ng recombinant SpGH92, respectively; lane 5: protein size ladder; lane 6–8: cell lysate from cells grown on mannose, glucose and galactose, respectively. No SpGH92 was detected in cell lysates; as a positive control, the same samples were blotted with an anti-GH20C antibody and GH20C was detected in the glucose-grown cell lysate as previously described [28]. (B) Screen of TIGR4 Sm r cellular fractions for SpGH92 activity by fluorophore-assisted carbohydrate electrophoresis (FACE).…”

Section: Supporting Informationmentioning

confidence: 99%

See 4 more Smart Citations

Molecular Characterization of N-glycan Degradation and Transport in Streptococcus pneumoniae and Its Contribution to Virulence

et al. 2017

Self Cite

View full text Add to dashboard Cite

The carbohydrate-rich coating of human tissues and cells provide a first point of contact for colonizing and invading bacteria. Commensurate with N-glycosylation being an abundant form of protein glycosylation that has critical functional roles in the host, some host-adapted bacteria possess the machinery to process N-linked glycans. The human pathogen Streptococcus pneumoniae depolymerizes complex N-glycans with enzymes that sequentially trim a complex N-glycan down to the Man3GlcNAc2 core prior to the release of the glycan from the protein by endo-β-N-acetylglucosaminidase (EndoD), which cleaves between the two GlcNAc residues. Here we examine the capacity of S. pneumoniae to process high-mannose N-glycans and transport the products. Through biochemical and structural analyses we demonstrate that S. pneumoniae also possesses an α-(1,2)-mannosidase (SpGH92). This enzyme has the ability to trim the terminal α-(1,2)-linked mannose residues of high-mannose N-glycans to generate Man5GlcNAc2. Through this activity SpGH92 is able to produce a substrate for EndoD, which is not active on high-mannose glycans with α-(1,2)-linked mannose residues. Binding studies and X-ray crystallography show that NgtS, the solute binding protein of an ABC transporter (ABCNG), is able to bind Man5GlcNAc, a product of EndoD activity, with high affinity. Finally, we evaluated the contribution of EndoD and ABCNG to growth of S. pneumoniae on a model N-glycosylated glycoprotein, and the contribution of these enzymes and SpGH92 to virulence in a mouse model. We found that both EndoD and ABCNG contribute to growth of S. pneumoniae, but that only SpGH92 and EndoD contribute to virulence. Therefore, N-glycan processing, but not transport of the released glycan, is required for full virulence in S. pneumoniae. To conclude, we synthesize our findings into a model of N-glycan processing by S. pneumoniae in which both complex and high-mannose N-glycans are targeted, and in which the two arms of this degradation pathway converge at ABCNG.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Supporting Informationmentioning

confidence: 99%

See 3 more Smart Citations

Molecular Characterization of N-glycan Degradation and Transport in Streptococcus pneumoniae and Its Contribution to Virulence

et al. 2017

Self Cite

View full text Add to dashboard Cite

show abstract

Glycan‐metabolizing enzymes in microbe–host interactions: the Streptococcus pneumoniae paradigm

2018

Self Cite

View full text Add to dashboard Cite

Streptococcus pneumoniae is a frequent colonizer of the upper airways; however, it is also an accomplished pathogen capable of causing life-threatening diseases. To colonize and cause invasive disease, this bacterium relies on a complex array of factors to mediate the host-bacterium interaction. The respiratory tract is rich in functionally important glycoconjugates that display a vast range of glycans, and, thus, a key component of the pneumococcus-host interaction involves an arsenal of bacterial carbohydrate-active enzymes to depolymerize these glycans and carbohydrate transporters to import the products. Through the destruction of host glycans, the glycan-specific metabolic machinery deployed by S. pneumoniae plays a variety of roles in the host-pathogen interaction. Here, we review the processing and metabolism of the major host-derived glycans, including N- and O-linked glycans, Lewis and blood group antigens, proteoglycans, and glycogen, as well as some dietary glycans. We discuss the role of these metabolic pathways in the S. pneumoniae-host interaction, speculate on the potential of key enzymes within these pathways as therapeutic targets, and relate S. pneumoniae as a model system to glycan processing in other microbial pathogens.

show abstract

Separation and Visualization of Glycans by Fluorophore-Assisted Carbohydrate Electrophoresis

Robb

Hobbs

Boraston

2023

Methods in Molecular Biology

View full text Add to dashboard Cite

A Second β-Hexosaminidase Encoded in the Streptococcus pneumoniae Genome Provides an Expanded Biochemical Ability to Degrade Host Glycans

Cited by 22 publications

References 53 publications

Molecular Characterization of N-glycan Degradation and Transport in Streptococcus pneumoniae and Its Contribution to Virulence

Molecular Characterization of N-glycan Degradation and Transport in Streptococcus pneumoniae and Its Contribution to Virulence

Glycan‐metabolizing enzymes in microbe–host interactions: the Streptococcus pneumoniae paradigm

Separation and Visualization of Glycans by Fluorophore-Assisted Carbohydrate Electrophoresis

Contact Info

Product

Resources

About