3-Isopropylmalate Dehydrogenase from an Acidophilic Chemolithotroph Thiobacillus ferrooxidans; Structure and Substrate Recognition
Kenji INAGAKI*a, Katsumi IMADAb, Hideyuki MATSUNAMIa, Hiroshi KAWAGUCHIa, Nobuo TANAKAc, Keiichi NAMBAb, and Hidehiko TANAKAa
a Department of Bioresources Chemistry, Okayama University, Faculty of Agriculture, 1-1-1 Tsushima-Naka, Okayama-Shi, Okayama 700, Japan
b International Institute for Advanced Research, Matsushita Electric Industrial Co., Ltd., 3-4 Hikaridai, Seika-Cho, Kyoto 619-02, Japan
c Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatuta, Midori-Ku, Yokohama-Shi, Kanagawa 226, Japan
3-Isopropylmalate dehydrogenase (IPMDH) is a dimeric enzyme that catalyzes dehydrogenation and decarboxylation reactions of IPM in the leucine biosynthetic pathway. IPMDH shows a marked similarity to isocitrate dehydrogenase (ICDH) in its structural framework and catalytic mechanism, and is classified into a unique group of decarboxylating dehydrogenases. Both enzymes catalyze the dehydrogenation at C2 and decarboxylation at C3 of (2R, 3S) 2-hydroxy acids and produce 2-keto acids. The difference in their substrates is the γ-moiety attached to 2R-malate, therefore these enzymes should recognize and distinguish the γ-moiety strictly. In fact, IPMDH demonstrates no activity on isocitrate, which is a substrate of ICDH, and vice versa. Recently we have determined the crystal structure of IPMDH from Thiobacillus ferrooxidans complexed with IPM at 2.0 A resolution by molecular replacement method. The structure actually shows a closed conformation and the substrate binding site is quite similar to the site of ICDH except for around theγ-isopropyl group. The γ-isopropyl group is recognized by a unique hydrophobic pocket which includes Glu88, Leu91, Leu92, and Va193'.
