title: Construction of a fully active truncated alternansucrase partially deleted of its carboxy-terminal domain.
abstract: Recombinant expression of the large alternansucrase (2057 amino acids) was hindered in E. coli due to poor enzyme solubility and protein degradation. The effects of deletions of the alternansucrase C-terminal CW-like and APY repeated motifs on enzyme solubility and specificity were investigated. A truncated variant deleted of the APY repeats but harboring four C-terminal CW-like repeats displayed a high specific activity and the same specificity of product synthesis as the native enzyme. It is more soluble and suffers less degradation than full length alternansucrase. Hence this truncated variant is a promising tool for the further structural and kinetic study of this interesting enzyme.
authors.affiliation: Ecole Supérieure de Technologie des Biomolécules de Bordeaux (ESTBB), Université Victor Segalen Bordeaux 2, 146 Rue Léo Saignat, 33076 Bordeaux Cedex, France.
authors.forename: Gilles
authors.initials: G
authors.lastname: Joucla
authors.forename: Sandra
authors.initials: S
authors.lastname: Pizzut
authors.forename: Pierre
authors.initials: P
authors.lastname: Monsan
authors.forename: Magali
authors.initials: M
authors.lastname: Remaud-Simeon
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D016695:Glycosyltransferases / Q000096:biosynthesis / Q000235:genetics*; D007934:Leuconostoc / Q000201:enzymology* / Q000235:genetics; D015202:Protein Engineering; D017434:Protein Structure, Tertiary / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D017384:Sequence Deletion; D013379:Substrate Specificity / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D016695:Glycosyltransferases; C033837:alternansucrase