Cell-associated alpha-amylases of butyrate-producing Firmicute bacteria from the human colon

Alan G. Ramsay, Karen Patricia Scott, Jennifer Cynthia Martin, Marco T. Rincon, Harry James Flint

Research output: Contribution to journalArticle

48 Citations (Scopus)

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 SIDS-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 center dot 6 kDa that includes a family 13 glycoside hydrolase domain, and duplicated family 26 carbohydrate-binding modules. Amy13A (182 center dot 4 kDa), from Roseburia inulinivorans A2-194, also includes a family 13 domain, which is preceded by two repeat units of similar to 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.

Original languageEnglish
Pages (from-to)3281-3290
Number of pages10
JournalMicrobiology
Volume152
DOIs
Publication statusPublished - Nov 2006

Keywords

  • carbohydrate-binding molecules
  • gram-positive bacteria
  • 16s ribosomal RNA
  • bacteroides thetaiotaomicron
  • human feces
  • molecular characterization
  • nucleotide sequence
  • starch utilization
  • resistant starch
  • short-chain

Cite this

Cell-associated alpha-amylases of butyrate-producing Firmicute bacteria from the human colon. / Ramsay, Alan G.; Scott, Karen Patricia; Martin, Jennifer Cynthia; Rincon, Marco T.; Flint, Harry James.

In: Microbiology , Vol. 152, 11.2006, p. 3281-3290.

Research output: Contribution to journalArticle

Ramsay, Alan G. ; Scott, Karen Patricia ; Martin, Jennifer Cynthia ; Rincon, Marco T. ; Flint, Harry James. / Cell-associated alpha-amylases of butyrate-producing Firmicute bacteria from the human colon. In: Microbiology . 2006 ; Vol. 152. pp. 3281-3290.
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AU - Flint, Harry James

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N2 - 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 SIDS-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 center dot 6 kDa that includes a family 13 glycoside hydrolase domain, and duplicated family 26 carbohydrate-binding modules. Amy13A (182 center dot 4 kDa), from Roseburia inulinivorans A2-194, also includes a family 13 domain, which is preceded by two repeat units of similar to 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.

AB - 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 SIDS-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 center dot 6 kDa that includes a family 13 glycoside hydrolase domain, and duplicated family 26 carbohydrate-binding modules. Amy13A (182 center dot 4 kDa), from Roseburia inulinivorans A2-194, also includes a family 13 domain, which is preceded by two repeat units of similar to 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.

KW - carbohydrate-binding molecules

KW - gram-positive bacteria

KW - 16s ribosomal RNA

KW - bacteroides thetaiotaomicron

KW - human feces

KW - molecular characterization

KW - nucleotide sequence

KW - starch utilization

KW - resistant starch

KW - short-chain

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DO - 10.1099/mic.0.29233-0

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JO - Microbiology

JF - Microbiology

SN - 1350-0872

ER -