title: Purification and characterization of NADP-linked isocitrate dehydrogenase from the copper-tolerant wood-rotting basidiomycete Fomitopsis palustris.
abstract: NADP-linked isocitrate dehydrogenase (EC 1.1.1.42), a key enzyme of the tricarboxylic acid cycle, was purified 672-fold as a nearly homogeneous protein from the copper-tolerant wood-rotting basidiomycete Fomitopsis palustris. The purified enzyme, with a molecular mass of 115 kDa, consisted of two 55-kDa subunits, and had the Km of 12.7, 2.9, and 23.9 microM for isocitrate, NADP, and Mg2+, respectively, at the optimal pH of 9.0. The enzyme had maximum activity in the presence of Mg2+, which also helped to prevent enzyme inactivation during the purification procedures and storage. The enzyme activity was competitively inhibited by 2-oxoglutarate (K(i), 127.0 microM). Although adenine nucleotides and other compounds, including some of the metabolic intermediates of glyoxylate and tricarboxylic acid cycles, had no or only slight inhibition, a mixture of oxaloacetate and glyoxylate potently inhibited the enzyme activity and the inhibition pattern was a mixed type.
authors.affiliation: Wood Research Institute, Kyoto University, Uji, Kyoto 611-0011, Japan.
authors.forename: Jeong-Jun
authors.initials: JJ
authors.lastname: Yoon
authors.forename: Takefumi
authors.initials: T
authors.lastname: Hattori
authors.forename: Mikio
authors.initials: M
authors.lastname: Shimada
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000249:Adenosine Monophosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001487:Basidiomycota / Q000378:metabolism*; D003300:Copper / Q000494:pharmacology*; D006038:Glyoxylates / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007521:Isocitrate Dehydrogenase / Q000737:chemistry* / Q000302:isolation & purification; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D010071:Oxaloacetates / Q000378:metabolism
chemical_list: D006038:Glyoxylates; D010071:Oxaloacetates; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D003300:Copper; D000255:Adenosine Triphosphate; D007521:Isocitrate Dehydrogenase; D008274:Magnesium; C031150:glyoxylic acid