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Structure of the GH76 α-mannanase homolog, BT2949, from the gut symbiont Bacteroides thetaiotaomicron.

Thompson AJ, Cuskin F, Spears RJ, Dabin J, Turkenburg JP, Gilbert HJ, Davies GJ - Acta Crystallogr. D Biol. Crystallogr. (2015)

Bottom Line: A significant member of this community, Bacteroides thetaiotaomicron, has evolved a complex system for sensing and processing a wide variety of natural glycoproducts in such a way as to provide maximum benefit to itself, the wider microbial community and the host.BT2949 presents a classical (α/α)6-barrel structure comprising a large extended surface cleft common to other GH76 family members.Analysis of the structure in conjunction with sequence alignments reveals the likely location of the catalytic active site of this noncanonical GH76.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, England.

ABSTRACT
The large bowel microbiota, a complex ecosystem resident within the gastrointestinal tract of all human beings and large mammals, functions as an essential, nonsomatic metabolic organ, hydrolysing complex dietary polysaccharides and modulating the host immune system to adequately tolerate ingested antigens. A significant member of this community, Bacteroides thetaiotaomicron, has evolved a complex system for sensing and processing a wide variety of natural glycoproducts in such a way as to provide maximum benefit to itself, the wider microbial community and the host. The immense ability of B. thetaiotaomicron as a `glycan specialist' resides in its enormous array of carbohydrate-active enzymes, many of which are arranged into polysaccharide-utilization loci (PULs) that are able to degrade sugar polymers that are often inaccessible to other gut residents, notably α-mannan. The B. thetaiotaomicron genome encodes ten putative α-mannanases spread across various PULs; however, little is known about the activity of these enzymes or the wider implications of α-mannan metabolism for the health of both the microbiota and the host. In this study, SAD phasing of a selenomethionine derivative has been used to investigate the structure of one such B. thetaiotaomicron enzyme, BT2949, which belongs to the GH76 family of α-mannanases. BT2949 presents a classical (α/α)6-barrel structure comprising a large extended surface cleft common to other GH76 family members. Analysis of the structure in conjunction with sequence alignments reveals the likely location of the catalytic active site of this noncanonical GH76.

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The structure of S. cerevisiae α-mannan and a potential model for its hydrolysis by Bt. Exo-acting GH92 α-­mannosidases remove α-1,2- and α-1,3-linked side-chain moieties, whilst GH76 enzymes hydrolyse the α-­1,6 backbone, allowing the import of smaller mono-oligosaccharides for further digestion. Carbohydrate residues are shown according to CFG (Centre for Functional Glycomics) nomenclature: green circles denote mannose, whilst blue squares represent N-acetylglucosamine. Linkages are specified.
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fig1: The structure of S. cerevisiae α-mannan and a potential model for its hydrolysis by Bt. Exo-acting GH92 α-­mannosidases remove α-1,2- and α-1,3-linked side-chain moieties, whilst GH76 enzymes hydrolyse the α-­1,6 backbone, allowing the import of smaller mono-oligosaccharides for further digestion. Carbohydrate residues are shown according to CFG (Centre for Functional Glycomics) nomenclature: green circles denote mannose, whilst blue squares represent N-acetylglucosamine. Linkages are specified.

Mentions: Bacteroides thetaiotaomicron (Bt), a dominant member of the microbiota, the symbiotic microbial community that inhabits the large bowel of humans and other higher mammals, plays an essential role in health and nutrition through its modulation of otherwise intractable dietary polysaccharides (Xu & Gordon, 2003 ▶; Bäckhed et al., 2005 ▶). Indeed, Bt represents one of the largest expansions of carbohydrate-processing enzymes known, with approximately 10% of its protein-encoding genes devoted solely to the production of catabolic glycoside hydrolase (GH) and anabolic glycosyltransferase (GT) enzymes (Xu et al., 2003 ▶). Of the various GHs encoded by Bt, a subset of over 30 enzymes are predicted to specifically target α-mannosidic linkages through their assignment to known α-mannosidase/α-mannanase families within the sequence-based CAZy classification (Lombard et al., 2014 ▶; http://www.cazy.org). The evolution of such a large catalytic repertoire to target a glycosidic bond observed only relatively scarcely in nature therefore suggests α-mannose-containing polysaccharides, such as the α-mannan present in yeast cell walls, to be a significant nutrient for both Bt and other closely related gut-dwelling bacteria. However, given their apparently low biological abundance (compared with, for example, β-mannose- or glucose-containing polysaccharides) and recalcitrant chemistry (nucleophilic substitution at the anomeric centre of mannosides is among the most difficult known; reviewed in Crich, 2010 ▶; Davies et al., 2011 ▶), the deliberate selection of α-mannosides as a primary carbon source therefore seems to be quite unusual. One likely line of reasoning for this utilization can be found in the continued evolution of the host diet. Over the last several thousand years, human beings have increasingly taken advantage of microorganism-catalysed fermentation reactions in the production of foods and beverages, such as bread and beer, a process exemplified by the now common referral to the budding yeast Saccharo­myces cerevisiae as so-called ‘brewer’s or baker’s yeast’. As such, consumption of yeast extracts, particularly α-mannan (for the structure of S. cerevisiae α-mannan, see Fig. 1 ▶), is now commonplace, strongly increasing their prevalence within the gut environment, and together with the highly competitive nature of the microbiota provides a strong evolutionary driving force towards the adaption of enzymes capable of processing this increasingly abundant and less commonly employed nutrient (Walter & Ley, 2011 ▶; Koropatkin et al., 2012 ▶).


Structure of the GH76 α-mannanase homolog, BT2949, from the gut symbiont Bacteroides thetaiotaomicron.

Thompson AJ, Cuskin F, Spears RJ, Dabin J, Turkenburg JP, Gilbert HJ, Davies GJ - Acta Crystallogr. D Biol. Crystallogr. (2015)

The structure of S. cerevisiae α-mannan and a potential model for its hydrolysis by Bt. Exo-acting GH92 α-­mannosidases remove α-1,2- and α-1,3-linked side-chain moieties, whilst GH76 enzymes hydrolyse the α-­1,6 backbone, allowing the import of smaller mono-oligosaccharides for further digestion. Carbohydrate residues are shown according to CFG (Centre for Functional Glycomics) nomenclature: green circles denote mannose, whilst blue squares represent N-acetylglucosamine. Linkages are specified.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4321491&req=5

fig1: The structure of S. cerevisiae α-mannan and a potential model for its hydrolysis by Bt. Exo-acting GH92 α-­mannosidases remove α-1,2- and α-1,3-linked side-chain moieties, whilst GH76 enzymes hydrolyse the α-­1,6 backbone, allowing the import of smaller mono-oligosaccharides for further digestion. Carbohydrate residues are shown according to CFG (Centre for Functional Glycomics) nomenclature: green circles denote mannose, whilst blue squares represent N-acetylglucosamine. Linkages are specified.
Mentions: Bacteroides thetaiotaomicron (Bt), a dominant member of the microbiota, the symbiotic microbial community that inhabits the large bowel of humans and other higher mammals, plays an essential role in health and nutrition through its modulation of otherwise intractable dietary polysaccharides (Xu & Gordon, 2003 ▶; Bäckhed et al., 2005 ▶). Indeed, Bt represents one of the largest expansions of carbohydrate-processing enzymes known, with approximately 10% of its protein-encoding genes devoted solely to the production of catabolic glycoside hydrolase (GH) and anabolic glycosyltransferase (GT) enzymes (Xu et al., 2003 ▶). Of the various GHs encoded by Bt, a subset of over 30 enzymes are predicted to specifically target α-mannosidic linkages through their assignment to known α-mannosidase/α-mannanase families within the sequence-based CAZy classification (Lombard et al., 2014 ▶; http://www.cazy.org). The evolution of such a large catalytic repertoire to target a glycosidic bond observed only relatively scarcely in nature therefore suggests α-mannose-containing polysaccharides, such as the α-mannan present in yeast cell walls, to be a significant nutrient for both Bt and other closely related gut-dwelling bacteria. However, given their apparently low biological abundance (compared with, for example, β-mannose- or glucose-containing polysaccharides) and recalcitrant chemistry (nucleophilic substitution at the anomeric centre of mannosides is among the most difficult known; reviewed in Crich, 2010 ▶; Davies et al., 2011 ▶), the deliberate selection of α-mannosides as a primary carbon source therefore seems to be quite unusual. One likely line of reasoning for this utilization can be found in the continued evolution of the host diet. Over the last several thousand years, human beings have increasingly taken advantage of microorganism-catalysed fermentation reactions in the production of foods and beverages, such as bread and beer, a process exemplified by the now common referral to the budding yeast Saccharo­myces cerevisiae as so-called ‘brewer’s or baker’s yeast’. As such, consumption of yeast extracts, particularly α-mannan (for the structure of S. cerevisiae α-mannan, see Fig. 1 ▶), is now commonplace, strongly increasing their prevalence within the gut environment, and together with the highly competitive nature of the microbiota provides a strong evolutionary driving force towards the adaption of enzymes capable of processing this increasingly abundant and less commonly employed nutrient (Walter & Ley, 2011 ▶; Koropatkin et al., 2012 ▶).

Bottom Line: A significant member of this community, Bacteroides thetaiotaomicron, has evolved a complex system for sensing and processing a wide variety of natural glycoproducts in such a way as to provide maximum benefit to itself, the wider microbial community and the host.BT2949 presents a classical (α/α)6-barrel structure comprising a large extended surface cleft common to other GH76 family members.Analysis of the structure in conjunction with sequence alignments reveals the likely location of the catalytic active site of this noncanonical GH76.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, England.

ABSTRACT
The large bowel microbiota, a complex ecosystem resident within the gastrointestinal tract of all human beings and large mammals, functions as an essential, nonsomatic metabolic organ, hydrolysing complex dietary polysaccharides and modulating the host immune system to adequately tolerate ingested antigens. A significant member of this community, Bacteroides thetaiotaomicron, has evolved a complex system for sensing and processing a wide variety of natural glycoproducts in such a way as to provide maximum benefit to itself, the wider microbial community and the host. The immense ability of B. thetaiotaomicron as a `glycan specialist' resides in its enormous array of carbohydrate-active enzymes, many of which are arranged into polysaccharide-utilization loci (PULs) that are able to degrade sugar polymers that are often inaccessible to other gut residents, notably α-mannan. The B. thetaiotaomicron genome encodes ten putative α-mannanases spread across various PULs; however, little is known about the activity of these enzymes or the wider implications of α-mannan metabolism for the health of both the microbiota and the host. In this study, SAD phasing of a selenomethionine derivative has been used to investigate the structure of one such B. thetaiotaomicron enzyme, BT2949, which belongs to the GH76 family of α-mannanases. BT2949 presents a classical (α/α)6-barrel structure comprising a large extended surface cleft common to other GH76 family members. Analysis of the structure in conjunction with sequence alignments reveals the likely location of the catalytic active site of this noncanonical GH76.

Show MeSH