Catalytic properties and crystal structure of UDP-galactose 4-epimerase-like l-threonine 3-dehydrogenase from Phytophthora infestans

• Published in: Enzyme and microbial technology Vol. 140; p. 109627
• Main Authors: Yoneda, Kazunari, Nagano, Rina, Mikami, Takuya, Sakuraba, Haruhiko, Fukui, Kenji, Araki, Tomohiro, Ohshima, Toshihisa
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.10.2020
• Subjects: 3-dehydrogenase; Alcohol Oxidoreductases - antagonists & inhibitors; Alcohol Oxidoreductases - chemistry; Alcohol Oxidoreductases - genetics; Alcohol Oxidoreductases - metabolism; Binding Sites; Catalysis; Catalytic Domain; Crystal structure; Crystallography, X-Ray; Enzyme Inhibitors - metabolism; Enzymological characterization; Glycine - analogs & derivatives; Glycine - metabolism; Hydrogen-Ion Concentration; Models, Molecular; Molecular Docking Simulation; Phytophthora infestans; Phytophthora infestans - enzymology; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Temperature; Threonine - metabolism; UDP-galactose 4-epimerase-like l-threonine; UDPglucose 4-Epimerase - antagonists & inhibitors; UDPglucose 4-Epimerase - chemistry; UDPglucose 4-Epimerase - genetics; UDPglucose 4-Epimerase - metabolism; SDR; Enzymological characterization; PEG; GalE-like l-ThrDH; Phytophthora infestans; 3-dehydrogenase; l-ThrDH; RMSD; UDP-galactose 4-epimerase-like l-threonine; Crystal structure
• ISSN: 0141-0229; 1879-0909
• DOI: 10.1016/j.enzmictec.2020.109627

•A UDP-galactose 4-epimerase-like l-threonine 3-dehydrogenase (GalE-like L-ThrDH) from Phytophthora infestans, a plant pathogenic fungus, was successfully isolated and characterized.•The N-acetylglycine, an inhibitor of P. infestans GalE-like L-ThrDH, is a potential lead compound to develop new highly specific agrochemicals against P. infestans.•The hexameric GalE-like L-ThrDH was determined using the molecular replacement method at a resolution of 2.3 Å, and the binding mode and inhibition mechanism of N-acetylglycine were elucidated. We report, for the first time, the three-dimensional structure and biochemical properties of a UDP-galactose 4-epimerase-like l-threonine 3-dehydrogenase (GalE-like L-ThrDH) from Phytophthora infestans, a plant disease-causing fungus. We identified GalE-like L-ThrDH using Kyoto Encyclopedia of Genes and Genomes (KEGG) database as a candidate target for the development of a new fungicide. The GalE-like L-ThrDH gene was expressed in Escherichia coli, and its product was purified and characterized. N-Acetylglycine was found to act as a competitive inhibitor of the enzyme (Ki =0.18 mM). The crystal structure of the unique hexameric GalE-like L-ThrDH was determined using the molecular replacement method at a resolution of 2.3 Å, in the presence of NAD+ and citrate, an analogue of the substrate. Based on the molecular docking simulation, N-acetylglycine molecule was modeled into the active site and the binding mode and inhibition mechanism of N-acetylglycine were elucidated.