title: Identification of sucrose synthase in nonphotosynthetic bacteria and characterization of the recombinant enzymes.
abstract: Sucrose synthase (SuSy) catalyzes the reversible conversion of sucrose and a nucleoside diphosphate into fructose and nucleotide (NDP)-glucose. To date, only SuSy's from plants and cyanobacteria, both photosynthetic organisms, have been characterized. Here, four prokaryotic SuSy enzymes from the nonphotosynthetic organisms Nitrosomonas Europaea (SuSyNe), Acidithiobacillus caldus (SuSyAc), Denitrovibrio acetiphilus (SusyDa), and Melioribacter roseus (SuSyMr) were recombinantly expressed in Escherichia coli and thoroughly characterized. The purified enzymes were found to display high-temperature optima (up to 80 °C), high activities (up to 125 U/mg), and high thermostability (up to 15 min at 60 °C). Furthermore, SuSyAc, SuSyNe, and SuSyDa showed a clear preference for ADP as nucleotide, as opposed to plant SuSy's which prefer UDP. A structural and mutational analysis was performed to elucidate the difference in NDP preference between eukaryotic and prokaryotic SuSy's. Finally, the physiological relevance of this enzyme specificity is discussed in the context of metabolic pathways and genomic organization.
authors.affiliation: Centre for Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure Links 653, 9000, Ghent, Belgium.
authors.forename: Margo
authors.initials: M
authors.lastname: Diricks
authors.affiliation: Centre for Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure Links 653, 9000, Ghent, Belgium.
authors.forename: Frederik
authors.initials: F
authors.lastname: De Bruyn
authors.affiliation: Centre for Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure Links 653, 9000, Ghent, Belgium.
authors.forename: Paul
authors.initials: P
authors.lastname: Van Daele
authors.affiliation: Centre for Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure Links 653, 9000, Ghent, Belgium.
authors.forename: Maarten
authors.initials: M
authors.lastname: Walmagh
authors.affiliation: Centre for Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure Links 653, 9000, Ghent, Belgium. tom.desmet@ugent.be.
authors.forename: Tom
authors.initials: T
authors.lastname: Desmet
mesh_terms: D001419:Bacteria / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D016000:Cluster Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004795:Enzyme Stability; D015870:Gene Expression; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D004269:DNA, Bacterial; D011994:Recombinant Proteins; D005964:Glucosyltransferases; C022699:sucrose synthase
keywords: Nucleotide sugar; Photosynthesis; Sucrose metabolism; Sucrose synthase