title: Characterization of D-xylonate dehydratase YjhG from Escherichia coli.
abstract: D-xylonate dehydratase YjhG from Escherichia coli can convert D-xylonate into 2-keto-3-deoxy- D-xylonate (KDX), and is a key enzyme in the biosynthesis of 1,2,4-butanetriol and other chemicals. However, the biochemical properties of YjhG still remain unknown. In this study, the activity assay method for YjhG was established based on semicarbazide method, in which KDX reacts with semicarbazide reagent, and is further quantified by high-resolution mass spectrometry. The effect of reaction conditions on YjhG activity was determined in vitro using purified His-tagged YjhG protein. This enzyme showed maximal activity at 30°C and pH 8.0. Bivalent metal ions such as Mg(2+) and Mn(2+) activated, whereas Ni(2+) and Zn(2+) inhibited the activity of YjhG. Under optimal conditions, the Km and Vmax values were 4.88 mM and 78.62 μM l(-1)h(-1), respectively, when using D-xylonate as a substrate. Amino acids sequence alignments and catalytic properties analysis revealed that YjhG might be a member of IlvD/EDD family. Results obtained in this study may lay a foundation for further investigation on YjhG and will benefit its application in biosynthesis of related chemicals.
authors.affiliation: a CAS Key Laboratory of Bio-based Materials; Qingdao Institute of Bioenergy and Bioprocess Technology; Chinese Academy of Sciences ; Qingdao , China.
authors.forename: Yudong
authors.initials: Y
authors.lastname: Jiang
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.forename: Tao
authors.initials: T
authors.lastname: Cheng
authors.forename: Yujin
authors.initials: Y
authors.lastname: Cao
authors.forename: Rubing
authors.initials: R
authors.lastname: Zhang
authors.forename: Mo
authors.initials: M
authors.lastname: Xian
mesh_terms: D000595:Amino Acid Sequence; D055162:Biocatalysis; D000440:Butanols / Q000378:metabolism; D057075:Enzyme Assays; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006836:Hydro-Lyases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013400:Sugar Acids / Q000378:metabolism; D013696:Temperature; D014994:Xylose / Q000378:metabolism
chemical_list: C583633:2-keto-3-deoxy-D-xylonate; D000440:Butanols; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D013400:Sugar Acids; C092647:lyxonic acid; D014994:Xylose; C037004:D-xylo-aldonate dehydratase; D006836:Hydro-Lyases; C114589:1,3,4-butanetriol
keywords: 1,2,4-butanetriol; D-xylonate dehydratase; Escherichia coli; YjhG; biosynthesis