title: Biosynthesis of nucleotide sugars by a promiscuous UDP-sugar pyrophosphorylase from Arabidopsis thaliana (AtUSP).
abstract: Nucleotide sugars are activated forms of monosaccharides and key intermediates of carbohydrate metabolism in all organisms. The availability of structurally diverse nucleotide sugars is particularly important for the characterization of glycosyltransferases. Given that limited methods are available for preparation of nucleotide sugars, especially their useful non-natural derivatives, we introduced herein an efficient one-step three-enzyme catalytic system for the synthesis of nucleotide sugars from monosaccharides. In this study, a promiscuous UDP-sugar pyrophosphorylase (USP) from Arabidopsis thaliana (AtUSP) was used with a galactokinase from Streptococcus pneumoniae TIGR4 (SpGalK) and an inorganic pyrophosphatase (PPase) to effectively synthesize four UDP-sugars. AtUSP has better tolerance for C4-derivatives of Gal-1-P compared to UDP-glucose pyrophosphorylase from S. pneumoniae TIGR4 (SpGalU). Besides, the nucleotide substrate specificity and kinetic parameters of AtUSP were systematically studied. AtUSP exhibited considerable activity toward UTP, dUTP and dTTP, the yield of which was 87%, 85% and 84%, respectively. These results provide abundant information for better understanding of the relationship between substrate specificity and structural features of AtUSP.
authors.affiliation: The State Key Laboratory of Microbial Technology and School of Life Sciences, Jinan, Shandong 250100, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
authors.forename: Yang
authors.initials: Y
authors.lastname: Zou
authors.forename: Wanyi
authors.initials: W
authors.lastname: Guan
authors.forename: Yafei
authors.initials: Y
authors.lastname: Zhai
authors.forename: Mengyang
authors.initials: M
authors.lastname: Xue
authors.forename: Lan
authors.initials: L
authors.lastname: Jin
authors.forename: Xueer
authors.initials: X
authors.lastname: Zhao
authors.forename: Junkai
authors.initials: J
authors.lastname: Dong
authors.forename: Wenjun
authors.initials: W
authors.lastname: Wang
authors.forename: Jie
authors.initials: J
authors.lastname: Shen
authors.forename: Peng George
authors.initials: PG
authors.lastname: Wang
authors.forename: Min
authors.initials: M
authors.lastname: Chen
mesh_terms: D017360:Arabidopsis / Q000201:enzymology* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000378:metabolism*; D002236:Carbohydrate Conformation; D005686:Galactokinase / Q000378:metabolism; D009713:Nucleotidyltransferases / Q000378:metabolism*; D011755:Pyrophosphatases / Q000378:metabolism; D013296:Streptococcus pneumoniae / Q000201:enzymology; D014539:Uridine Diphosphate Sugars / Q000096:biosynthesis* / Q000737:chemistry
chemical_list: D029681:Arabidopsis Proteins; D014539:Uridine Diphosphate Sugars; D005686:Galactokinase; D009713:Nucleotidyltransferases; C514559:UDP-sugar pyrophosphorylase, Arabidopsis; D011755:Pyrophosphatases