期刊
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
卷 585, 期 -, 页码 39-51出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2015.08.025
关键词
Crystal structure; UDP-galactose 4-epimerase; Substrate specificity; Hyperthermophiles; Evolution
资金
- National Research Foundation of Korea (NRF) - Korean government (MSIP) [2014R1A2A2A01006765, 2013R1A1A2057465]
- Korea Institute of Planning & Evaluation for Technology (iPET) - Ministry for Food, Agriculture, Forestry and Fisheries [311042-05-1-HD120]
- National Research Foundation of Korea [2014R1A2A2A01006765, 2013R1A1A2057465] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
UDP-galactose 4-epimerase (GalE) catalyzes the interconversion of UDP-glucose (UDP-Glc) and UDPgalactose (UDP-Gal), which is a pivotal step in the Leloir pathway for D-galactose metabolism. Although GalE is widely distributed in prokaryotes and eukaryotes, little information is available regarding hyperthermophilic GalE. We overexpressed the TM0509 gene, encoding a putative GalE from Thermotoga maritima (TMGalE), in Escherichia coli and characterized the encoded protein. To further investigate the molecular basis of this enzyme's catalytic function, we determined the crystal structures of TMGalE and TMGalE bound to UDP-Glc at resolutions of 1.9 angstrom and 2.0 angstrom, respectively. The enzyme was determined to be a homodimer with a molecular mass of 70 kDa. The enzyme could reversibly catalyze the epimerization of UDP-GalNAc/UDP-GlcNAc as well as UDP-Gal/UDP-Glc at elevated temperatures, with an apparent optimal temperature and pH of 80 degrees C and 7.0, respectively. Our data showed that TM0509 is a UDP-galactosugar 4-epimerase involved in D-galactose metabolism; consequently, this study provides the first detailed characterization of a hyperthermophilic GalE. Moreover, the promiscuous substrate specificity of TMGalE, which is more similar to human GalE than E. coli GalE, supports the notion that TMGalE might exhibit the earliest form of sugar-epimerizing enzymes in the evolution of galactose metabolism. (C) 2015 Elsevier Inc. All rights reserved.
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