Dataset Information

Characterization of a human core-specific lysosomal {alpha}1,6-mannosidase involved in N-glycan catabolism.

ABSTRACT: In humans and rodents, the lysosomal catabolism of core Man(3)GlcNAc(2) N-glycan structures is catalyzed by the concerted action of several exoglycosidases, including a broad specificity lysosomal alpha-mannosidase (LysMan), core-specific alpha1,6-mannosidase, beta-mannosidase, and cleavage at the reducing terminus by a di-N-acetylchitobiase. We describe here the first cloning, expression, purification, and characterization of a novel human glycosylhydrolase family 38 alpha-mannosidase with catalytic characteristics similar to those established previously for the core-specific alpha1,6-mannosidase (acidic pH optimum, inhibition by swainsonine and 1,4-dideoxy-1,4-imino-d-mannitol, and stimulation by Co(2+) and Zn(2+)). Substrate specificity studies comparing the novel human alpha-mannosidase with human LysMan revealed that the former enzyme efficiently cleaved only the alpha1-6mannose residue from Man(3)GlcNAc but not Man(3)GlcNAc(2) or other larger high mannose oligosaccharides, indicating a requirement for chitobiase action before alpha1,6-mannosidase activity. In contrast, LysMan cleaved all of the alpha-linked mannose residues from high mannose oligosaccharides except the core alpha1-6mannose residue. alpha1,6-Mannosidase transcripts were ubiquitously expressed in human tissues, and expressed sequence tag searches identified homologous sequences in murine, porcine, and canine databases. No expressed sequence tags were identified for bovine alpha1,6-mannosidase, despite the identification of two sequence homologs in the bovine genome. The lack of conservation in 5'-flanking sequences for the bovine alpha1,6-mannosidase genes may lead to defective transcription similar to transcription defects in the bovine chitobiase gene. These results suggest that the chitobiase and alpha1,6-mannosidase function in tandem for mammalian lysosomal N-glycan catabolism.

SUBMITTER: Park C

PROVIDER: S-EPMC1351102 | biostudies-literature | 2005 Nov

REPOSITORIES: biostudies-literature

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Characterization of a human core-specific lysosomal {alpha}1,6-mannosidase involved in N-glycan catabolism.

Park Chaeho C Meng Lu L Stanton Leslie H LH Collins Robert E RE Mast Steven W SW Yi Xiaobing X Strachan Heather H Moremen Kelley W KW

The Journal of biological chemistry 20050822 44

In humans and rodents, the lysosomal catabolism of core Man(3)GlcNAc(2) N-glycan structures is catalyzed by the concerted action of several exoglycosidases, including a broad specificity lysosomal alpha-mannosidase (LysMan), core-specific alpha1,6-mannosidase, beta-mannosidase, and cleavage at the reducing terminus by a di-N-acetylchitobiase. We describe here the first cloning, expression, purification, and characterization of a novel human glycosylhydrolase family 38 alpha-mannosidase with cata ...[more]

PMID: 16115860

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Project description:The survival of commensal bacteria in the human gut partially depends on their ability to metabolize host-derived molecules. The use of the glycosidic moiety of N-glycoproteins by bacteria has been reported, but the role of N-glycopeptides or glycoamino acids as the substrates for bacterial growth has not been evaluated. We have identified in Lactobacillus casei strain BL23 a gene cluster (alf-2) involved in the catabolism of the glycoamino acid fucosyl-α-1,6-N-GlcNAc-Asn (6'FN-Asn), a constituent of the core-fucosylated structures of mammalian N-glycoproteins. The cluster consists of the genes alfHC, encoding a major facilitator superfamily (MFS) permease and the α-l-fucosidase AlfC, and the divergently oriented asdA (aspartate 4-decarboxylase), alfR2 (transcriptional regulator), pepV (peptidase), asnA2 (glycosyl-asparaginase), and sugK (sugar kinase) genes. Knockout mutants showed that alfH, alfC, asdA, asnA2, and sugK are necessary for efficient 6'FN-Asn utilization. The alf-2 genes are induced by 6'FN-Asn, but not by its glycan moiety, via the AlfR2 regulator. The constitutive expression of alf-2 genes in an alfR2 strain allowed the metabolism of a variety of 6'-fucosyl-glycans. However, GlcNAc-Asn did not support growth in this mutant background, indicating that the presence of a 6'-fucose moiety is crucial for substrate transport via AlfH. Within bacteria, 6'FN-Asn is defucosylated by AlfC, generating GlcNAc-Asn. This glycoamino acid is processed by the glycosylasparaginase AsnA2. GlcNAc-Asn hydrolysis generates aspartate and GlcNAc, which is used as a fermentable source by L.casei These data establish the existence in a commensal bacterial species of an exclusive metabolic pathway likely to scavenge human milk and mucosal fucosylated N-glycopeptides in the gastrointestinal tract.IMPORTANCE The gastrointestinal tract accommodates more than 1014 microorganisms that have an enormous impact on human health. The mechanisms enabling commensal bacteria and administered probiotics to colonize the gut remain largely unknown. The ability to utilize host-derived carbon and energy resources available at the mucosal surfaces may provide these bacteria with a competitive advantage in the gut. Here, we have identified in the commensal species Lactobacillus casei a novel metabolic pathway for the utilization of the glycoamino acid fucosyl-α-1,6-N-GlcNAc-Asn, which is present in the core-fucosylated N-glycoproteins from mammalians. These results give insight into the molecular interactions between the host and commensal/probiotic bacteria and may help to devise new strategies to restore gut microbiota homeostasis in diseases associated with dysbiotic microbiota.

Dataset Information

Characterization of a human core-specific lysosomal {alpha}1,6-mannosidase involved in N-glycan catabolism.

Publications

Characterization of a human core-specific lysosomal {alpha}1,6-mannosidase involved in N-glycan catabolism.

Similar Datasets

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