title: Noncompetitive inhibition by L-cysteine and activation by L-glutamate of the iron-oxidizing activity of a mixotrophic iron-oxidizing bacterium strain OKM-9.
abstract: A mesophilic, mixotrophic iron-oxidizing bacterium strain OKM-9 uses ferrous iron as a sole source of energy and L-glutamate as a sole source of cellular carbon. Uptake of L-glutamate into OKM-9 cells is absolutely dependent on ferrous iron oxidation. Thus, the Fe(2+)-dependent L-glutamate uptake system of strain OKM-9 is crucial for the bacterium to grow mixotrophically in iron medium with L-glutamate. The relationship between iron oxidation and L-glutamate transport activities was studied. Iron oxidase containing cytochrome a was purified 9-fold from the plasma membrane of OKM-9. A purified iron oxidase showed one rust-colored band following disc gel electrophoresis after incubation with Fe(2+). The Fe(2+)-dependent L-glutamate transport system was also purified 14.5-fold from the plasma membrane using the same purification steps as for iron oxidase. Fe(2+)-dependent L-glutamate and L-cysteine uptake activities of OKM-9 were 0.36 and 0.24 nmol/mg/min, respectively, when a concentration of 18 mM of these amino acids was used as a substrate. Both uptake activities were completely inhibited by potassium cyanide (KCN), suggesting that cytochrome a in the iron oxidase is involved in the transport process. The iron-oxidizing activity of strain OKM-9 was activated 1.7-fold by 80 mM L-glutamate. In contrast, the activity was noncompetitively inhibited by L-cysteine. The Michaelis constant of iron oxidase for Fe(2+) was 12.6 mM and the inhibition constant for L-cysteine was 41.6 mM. A marked inhibition of iron oxidase by 50 mM L-cysteine was completely reversed by the addition of 60 mM L-glutamate. The results suggest the possibility that iron oxidase has a binding site for L-cysteine and the cysteine first bound to the iron oxidase was replaced by the added L-glutamate.
authors.affiliation: Division of Science and Technology for Energy Conversion, Graduate School of Natural Science and Technology, Okayama University 3-1-1, Tsushima-naka, Okayama, 700-8530, Japan. sugio@cc.okayama-u.ac.jp
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Sugio
authors.forename: Takao
authors.initials: T
authors.lastname: Inoue
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Kitano
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Takeuchi
authors.forename: Kazuo
authors.initials: K
authors.lastname: Kamimura