title: Mutational analysis of the role of calcium ions in the Lactobacillus reuteri strain 121 fructosyltransferase (levansucrase and inulosucrase) enzymes.
abstract: Bacterial fructosyltransferase enzymes belonging to glycoside hydrolase family 68 (GH68) are not known to require a metal cofactor. Here, we show that Ca2+ ions play an important structural role in the Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes. Analysis of the Bacillus subtilis Lev 3D structure [Meng, G. and Futterer, K. (2003) Nat. Struct. Biol. 10, 935-941] has provided evidence for the presence of a bound metal ion, most likely Ca2+. Characterization of site-directed mutants in the putative Ca2+ ion-binding sites of Lb. reuteri Lev and Inu revealed that the Inu Asp520 and Lev Asp500 residues play an important role in Ca2+ binding. Sequence alignments of family GH68 proteins showed that this Ca2+ ion-binding site is (largely) present only in proteins of Gram-positive origin.
authors.affiliation: Centre for Carbohydrate Bioengineering, TNO-University of Groningen, PO Box 14, 9750 AA Haren, The Netherlands.
authors.forename: L K
authors.initials: LK
authors.lastname: Ozimek
authors.forename: G J W
authors.initials: GJ
authors.lastname: Euverink
authors.forename: M J E C
authors.initials: MJ
authors.lastname: van der Maarel
authors.forename: L
authors.initials: L
authors.lastname: Dijkhuizen
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D004492:Edetic Acid / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D006602:Hexosyltransferases / Q000737:chemistry / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D017136:Ion Transport; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D011489:Protein Denaturation / Q000187:drug effects; D016415:Sequence Alignment; D013696:Temperature; D001426:Bacterial Proteins
chemical_list: D002118:Calcium; D004492:Edetic Acid; D006602:Hexosyltransferases; C021513:levansucrase; C021451:inulosucrase; D001426:Bacterial Proteins