title: Cell-associated alpha-amylases of butyrate-producing Firmicute bacteria from the human colon.
abstract: Selected butyrate-producing bacteria from the human colon that are related to Roseburia spp. and Butyrivibrio fibrisolvens showed a good ability to utilize a variety of starches for growth when compared with the Gram-negative amylolytic anaerobe Bacteroides thetaiotaomicron. A major cell-associated amylase of high molecular mass (140-210 kDa) was detected in each strain by SDS-PAGE zymogram analysis, and genes corresponding to these enzymes were analysed for two representative strains. Amy13B from But. fibrisolvens 16/4 is a multi-domain enzyme of 144.6 kDa that includes a family 13 glycoside hydrolase domain, and duplicated family 26 carbohydrate-binding modules. Amy13A (182.4 kDa), from Roseburia inulinivorans A2-194, also includes a family 13 domain, which is preceded by two repeat units of approximately 116 aa rich in aromatic residues, an isoamylase N-terminal domain, a pullulanase-associated domain, and an additional unidentified domain. Both Amy13A and Amy13B have N-terminal signal peptides and C-terminal cell-wall sorting signals, including a modified LPXTG motif similar to that involved in interactions with the cell surface in other Gram-positive bacteria, a hydrophobic transmembrane segment, and a basic C terminus. The overexpressed family 13 domains showed an absolute requirement for Mg2+ or Ca2+ for activity, and functioned as 1,4-alpha-glucanohydrolases (alpha-amylases; EC 3.2.1.1). These major starch-degrading enzymes thus appear to be anchored to the cell wall in this important group of human gut bacteria.
authors.affiliation: Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, UK.
authors.forename: Alan G
authors.initials: AG
authors.lastname: Ramsay
authors.affiliation: Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, UK.
authors.forename: Karen P
authors.initials: KP
authors.lastname: Scott
authors.affiliation: Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, UK.
authors.forename: Jenny C
authors.initials: JC
authors.lastname: Martin
authors.affiliation: Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, UK.
authors.forename: Marco T
authors.initials: MT
authors.lastname: Rincon
authors.affiliation: Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB, UK.
authors.forename: Harry J
authors.initials: HJ
authors.lastname: Flint
mesh_terms: D001421:Bacteria, Anaerobic / Q000145:classification / Q000201:enzymology* / Q000378:metabolism; D002087:Butyrates / Q000378:metabolism*; D002473:Cell Wall / Q000201:enzymology*; D003106:Colon / Q000382:microbiology*; D005798:Genes, Bacterial; D006801:Humans; D006867:Hydrolases / Q000145:classification / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D017434:Protein Structure, Tertiary; D013213:Starch / Q000378:metabolism; D000516:alpha-Amylases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D002087:Butyrates; D013213:Starch; D006867:Hydrolases; D000516:alpha-Amylases