title: Characterization and expression of glucosamine-6-phosphate synthase from Saccharomyces cerevisiae in Pichia pastoris.
abstract: Glucosamine-6-phosphate (GlcN-6-P) synthase from Saccharomyces cerevisiae was expressed in Pichia pastoris SMD1168 GIVING maximum activity of 96 U ml(-1) for the enzyme in the culture medium. By SDS-PAGE, the enzyme, a glycosylated protein, had an apparent molecular mass of 90 kDa. The enzyme was purified by gel exclusion chromatography to near homogeneity, with a 90 % yield and its properties were characterized. Optimal activities were at pH 5.5 and 55 °C, respectively, at which the highest specific activity was 6.8 U mg protein (-1). The enzyme was stable from pH 4.5 to 5.5 and from 45 to 60 °C. The Km and Vmax of the GlcN-6-P synthase towards D-fructose 6-phosphate were 2.8 mM and 6.9 μmol min(-1) mg(-1), respectively.
authors.affiliation: Shandong Provincial Key Laboratory of Microbial Engineering, School of Food and Bioengineering, Qilu University of Technology, Jinan, 250353, China, wangshengshow@sina.com.
authors.forename: Sheng
authors.initials: S
authors.lastname: Wang
authors.forename: Piwu
authors.initials: P
authors.lastname: Li
authors.forename: Jing
authors.initials: J
authors.lastname: Su
authors.forename: Xiangkun
authors.initials: X
authors.lastname: Wu
authors.forename: Rongrong
authors.initials: R
authors.lastname: Liang
mesh_terms: D003001:Cloning, Molecular; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing) / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D008958:Models, Molecular; D010843:Pichia / Q000201:enzymology / Q000235:genetics*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C098848:GFA1 protein, S cerevisiae; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)