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PUL0029 |
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30564851, Appl Microbiol Biotechnol. 2019 Feb;103(3):1299-1310. doi: 10.1007/s00253-018-9566-4. Epub 2018 Dec 18. |
| | Degradative enzymes for type II arabinogalactan side chains in Bifidobacterium longum subsp. longum. |
| | Fujita K, Sakamoto A, Kaneko S, Kotake T, Tsumuraya Y, Kitahara K |
| | Type II arabinogalactan (AG) is a soluble prebiotic fiber stimulating the proliferation of bifidobacteria in the human gut. Larch AG, which is comprised of type II AG, is known to be utilized as an energy source for Bifidobacterium longum subsp. longum (B. longum). We have previously characterized GH43_24 exo-beta-1,3-galactanase (Bl1,3Gal) for the degradation of type II AG main chains in B. longum JCM1217. In this study, we characterized GH30_5 exo-beta-1,6-galactobiohydrolase (Bl1,6Gal) and GH43_22 alpha-L-arabinofuranosidase (BlArafA), which are degradative enzymes for type II AG side chains in cooperation with exo-beta-1,3-galactanase. The recombinant exo-beta-1,6-galactobiohydrolase specifically released beta-1,6-galactobiose (beta-1,6-Gal(2)) from the nonreducing terminal of beta-1,6-galactooligosaccharides, and the recombinant alpha-L-arabinofuranosidase released arabinofuranose (Araf) from alpha-1,3-Araf-substituted beta-1,6-galactooligosaccharides. beta-1,6-Gal(2) was additively released from larch AG by the combined use of type II AG degradative enzymes, including Bl1,3Gal, Bl1,6Gal, and BlArafA. The gene cluster encoding the type II AG degradative enzymes is conserved in all B. longum strains, but not in other bifidobacterial species. The degradative enzymes for type II AG side chains are thought to be important for the acquisition of type II AG in B. longum. |
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30635377, Appl Environ Microbiol. 2019 Mar 6;85(6):e02582-18. doi: 10.1128/AEM.02582-18. Print 2019 Mar 15. |
| | Two Novel alpha-l-Arabinofuranosidases from Bifidobacterium longum subsp. longum Belonging to Glycoside Hydrolase Family 43 Cooperatively Degrade Arabinan. |
| | Komeno M, Hayamizu H, Fujita K, Ashida H |
| | 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 alpha-l-arabinofuranosidase activity toward p-nitrophenyl-alpha-l-arabinofuranoside but no activity toward p-nitrophenyl-beta-d-xylopyranoside. The specificities of the two enzymes for l-arabinofuranosyl linkages were different. BlArafC catalyzed the hydrolysis of alpha1,2- and alpha1,3-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 alpha1,5-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 alpha-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 alpha1,2/3-l-arabinofuranosyl linkages, while the latter acts on the alpha1,5-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. |
