Biosynthesis of nucleotide sugars by a promiscuous UDP-sugar pyrophosphorylase from Arabidopsis thaliana (AtUSP)

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...

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Bibliographic Details
Published in:Bioorganic & medicinal chemistry letters 2013-07, Vol.23 (13), p.3764-3768
Main Authors: Liu, Jun, Zou, Yang, Guan, Wanyi, Zhai, Yafei, Xue, Mengyang, Jin, Lan, Zhao, Xueer, Dong, Junkai, Wang, Wenjun, Shen, Jie, Wang, Peng George, Chen, Min
Format: Article
Language:English
Subjects:
Arabidopsis - enzymology
Arabidopsis - metabolism
Arabidopsis Proteins - metabolism
Arabidopsis thaliana
Carbohydrate Conformation
Galactokinase
Galactokinase - metabolism
Nucleotide sugars
Nucleotidyltransferases - metabolism
One-pot
Pyrophosphatases - metabolism
Streptococcus pneumoniae
Streptococcus pneumoniae - enzymology
UDP-sugar pyrophosphorylase
Uridine Diphosphate Sugars - biosynthesis
Uridine Diphosphate Sugars - chemistry
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Summary: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.
ISSN:0960-894X
1464-3405
DOI:10.1016/j.bmcl.2013.04.090