title: Molecular insights into the mechanism of thermal stability of actinomycete mannanase.
abstract: Streptomyces thermolilacinus mannanase (StMan), which requires Ca(2+) for its enhanced thermal stability and hydrolysis activity, possesses two Ca(2+) -binding sites in loop6 and loop7. We evaluated the function of the Ca(2+) -binding site in loop7 and the hydrogen bond between residues Ser247 in loop6 and Asp279 in loop7. The Ca(2+) -binding in loop7 was involved only in thermal stability. Mutations of Ser247 or Asp279 retained the Ca(2+) -binding ability; however, mutants showed less thermal stability than StMan. Phylogenetic analysis indicated that most glycoside hydrolase family 5 subfamily 8 mannanases could be stabilized by Ca(2+) ; however, the mechanism of StMan thermal stability was found to be quite specific in some actinomycete mannanases.
authors.affiliation: Okayama Prefectural Technology Center for Agriculture, Forestry and Fisheries, Research Institute for Biological Sciences (RIBS), Okayama, Japan.|Research Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
authors.forename: Yuya
authors.initials: Y
authors.lastname: Kumagai
authors.affiliation: Okayama Prefectural Technology Center for Agriculture, Forestry and Fisheries, Research Institute for Biological Sciences (RIBS), Okayama, Japan.
authors.forename: Misugi
authors.initials: M
authors.lastname: Uraji
authors.affiliation: Okayama Prefectural Technology Center for Agriculture, Forestry and Fisheries, Research Institute for Biological Sciences (RIBS), Okayama, Japan.
authors.forename: Kun
authors.initials: K
authors.lastname: Wan
authors.affiliation: Research Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
authors.forename: Masayuki
authors.initials: M
authors.lastname: Okuyama
authors.affiliation: Research Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
authors.forename: Atsuo
authors.initials: A
authors.lastname: Kimura
authors.affiliation: Okayama Prefectural Technology Center for Agriculture, Forestry and Fisheries, Research Institute for Biological Sciences (RIBS), Okayama, Japan.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Hatanaka
mesh_terms: D001665:Binding Sites; D002118:Calcium / Q000737:chemistry; D004795:Enzyme Stability; D006868:Hydrolysis; D010802:Phylogeny; D011487:Protein Conformation; D013302:Streptomyces / Q000737:chemistry / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D044902:beta-Mannosidase / Q000737:chemistry* / Q000378:metabolism
chemical_list: D044902:beta-Mannosidase; D002118:Calcium
keywords: actinomycete; glycoside hydrolase family 5; mannanase