Dataset Information

Two extracellular α-arabinofuranosidases are required for cereal-derived arabinoxylan metabolism by Bifidobacterium longum subsp. longum.

ABSTRACT: Members of the genus Bifidobacterium are commonly found in the human gut and are known to utilize complex carbohydrates that are indigestible by the human host. Members of the Bifidobacterium longum subsp. longum taxon can metabolize various plant-derived carbohydrates common to the human diet. To metabolize such polysaccharides, which include arabinoxylan, bifidobacteria need to encode appropriate carbohydrate-active enzymes in their genome. In the current study, we describe two GH43 family enzymes, denoted here as AxuA and AxuB, which are encoded by B. longum subsp. longum NCIMB 8809 and are shown to be required for cereal-derived arabinoxylan metabolism by this strain. Based on the observed hydrolytic activity of AxuA and AxuB, assessed by employing various synthetic and natural substrates, and based on in silico analyses, it is proposed that both AxuA and AxuB represent extracellular α-L-arabinofuranosidases with distinct substrate preferences. The variable presence of the axuA and axuB genes and other genes previously described to be involved in the metabolism of arabinose-containing glycans can in the majority cases explain the (in)ability of individual B. longum subsp. longum strains to grow on cereal-derived arabinoxylans and arabinan.

SUBMITTER: Friess L

PROVIDER: S-EPMC11318964 | biostudies-literature | 2024 Jan-Dec

REPOSITORIES: biostudies-literature

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Two extracellular α-arabinofuranosidases are required for cereal-derived arabinoxylan metabolism by Bifidobacterium longum subsp. longum.

Friess Lisa L Bottacini Francesca F McAuliffe Fionnuala M FM O'Neill Ian J IJ Cotter Paul D PD Lee Ciaran C Munoz-Munoz Jose J van Sinderen Douwe D

Gut microbes 20240101 1

Members of the genus Bifidobacterium are commonly found in the human gut and are known to utilize complex carbohydrates that are indigestible by the human host. Members of the Bifidobacterium longum subsp. longum taxon can metabolize various plant-derived carbohydrates common to the human diet. To metabolize such polysaccharides, which include arabinoxylan, bifidobacteria need to encode appropriate carbohydrate-active enzymes in their genome. In the current study, we describ ...[more]

PMID: 38752423

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Project description:Arabinose-containing poly- or oligosaccharides are suitable carbohydrate sources for Bifidobacterium longum subsp. longum However, their degradation pathways are poorly understood. In this study, we cloned and characterized the previously uncharacterized glycoside hydrolase family 43 (GH43) enzymes B. longum subsp. longum ArafC (BlArafC; encoded by BLLJ_1852) and B. longum subsp. longum ArafB (BlArafB; encoded by BLLJ_1853) from B. longum subsp. longum JCM 1217. Both enzymes exhibited α-l-arabinofuranosidase activity toward p-nitrophenyl-α-l-arabinofuranoside but no activity toward p-nitrophenyl-β-d-xylopyranoside. The specificities of the two enzymes for l-arabinofuranosyl linkages were different. BlArafC catalyzed the hydrolysis of α1,2- and α1,3-l-arabinofuranosyl linkages found on the side chains of both arabinan and arabinoxylan. It released l-arabinose 100 times faster from arabinan than from arabinoxylan but did not act on arabinogalactan. On the other hand, BlArafB catalyzed the hydrolysis of the α1,5-l-arabinofuranosyl linkage found on the arabinan backbone. It released l-arabinose from arabinan but not from arabinoxylan or arabinogalactan. Coincubation of BlArafC and BlArafB revealed that these two enzymes are able to degrade arabinan in a synergistic manner. Both enzyme activities were suppressed with EDTA treatment, suggesting that they require divalent metal ions. The GH43 domains of BlArafC and BlArafB are classified into GH43 subfamilies 27 and 22, respectively, but show very low similarity (less than 15% identity) with other biochemically characterized members in the corresponding subfamilies. The B. longum subsp. longum strain lacking the GH43 gene cluster that includes BLLJ_1850 to BLLJ_1853 did not grow in arabinan medium, suggesting that BlArafC and BlArafB are important for assimilation of arabinan.IMPORTANCE We identified two novel α-l-arabinofuranosidases, BlArafC and BlArafB, from B. longum subsp. longum JCM 1217, both of which are predicted to be extracellular membrane-bound enzymes. The former specifically acts on α1,2/3-l-arabinofuranosyl linkages, while the latter acts on the α1,5-l-arabinofuranosyl linkage. These enzymes cooperatively degrade arabinan and are required for the efficient growth of bifidobacteria in arabinan-containing medium. The genes encoding these enzymes are located side by side in a gene cluster involved in metabolic pathways for plant-derived polysaccharides, which may confer adaptability in adult intestines.

Project description:Gum arabic arabinogalactan (AG) protein (AGP) is a unique dietary fiber that is degraded and assimilated by only specific strains of Bifidobacterium longum subsp. longum Here, we identified a novel 3-O-α-d-galactosyl-α-l-arabinofuranosidase (GAfase) from B. longum JCM7052 and classified it into glycoside hydrolase family 39 (GH39). GAfase released α-d-Galp-(1→3)-l-Ara and β-l-Arap-(1→3)-l-Ara from gum arabic AGP and β-l-Arap-(1→3)-l-Ara from larch AGP, and the α-d-Galp-(1→3)-l-Ara release activity was found to be 594-fold higher than that of β-l-Arap-(1→3)-l-Ara. The GAfase gene was part of a gene cluster that included genes encoding a GH36 α-galactosidase candidate and ABC transporters for the assimilation of the released α-d-Galp-(1→3)-l-Ara in B. longum Notably, when α-d-Galp-(1→3)-l-Ara was removed from gum arabic AGP, it was assimilated by both B. longum JCM7052 and the nonassimilative B. longum JCM1217, suggesting that the removal of α-d-Galp-(1→3)-l-Ara from gum arabic AGP by GAfase permitted the cooperative action with type II AG degradative enzymes in B. longum The present study provides new insight into the mechanism of gum arabic AGP degradation in B. longumIMPORTANCE Bifidobacteria harbor numerous carbohydrate-active enzymes that degrade several dietary fibers in the gastrointestinal tract. B. longum JCM7052 is known to exhibit the ability to assimilate gum arabic AGP, but the key enzyme involved in the degradation of gum arabic AGP remains unidentified. Here, we cloned and characterized a GH39 3-O-α-d-galactosyl-α-l-arabinofuranosidase (GAfase) from B. longum JCM7052. The enzyme was responsible for the release of α-d-Galp-(1→3)-l-Ara and β-l-Arap-(1→3)-l-Ara from gum arabic AGP. The presence of a gene cluster including the GAfase gene is specifically observed in gum arabic AGP assimilative strains. However, GAfase carrier strains may affect GAfase noncarrier strains that express other type II AG degradative enzymes. These findings provide insights into the bifidogenic effect of gum arabic AGP.

Dataset Information

Two extracellular α-arabinofuranosidases are required for cereal-derived arabinoxylan metabolism by Bifidobacterium longum subsp. longum.

Publications

Two extracellular α-arabinofuranosidases are required for cereal-derived arabinoxylan metabolism by <i>Bifidobacterium longum</i> subsp. <i>longum</i>.

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