Purification and Characterization of Phosphoenolpyruvate Carboxylase of Photomixotrophically Cultured Green Tobacco Cells
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概要
- 論文の詳細を見る
Phosphoenolpyruvate (PEP) carboxylase (PEPCase, EC 4.1.1.31) was purified to apparent electrophoretic homogeneity from photomixotrophically cultured tobacco cells by ammonium sulfate fractionation, DEAE-Sephacel-, hydroxylapatite-, Phenyl-Sepharose CL-4B-, and Sepharose CL-6B-chromatography; and fast protein liquid chromatography on Mono Q. The purified enzyme had a specific activity of 32 units per mg protein, and its purity was determined by denaturing polyacrylamide gel electrophoresis. The native enzyme, with a molecular weight of about 440,000, was a tetramer of four identical subunits and showed maximum activity at pH 8.5-9.0. Non-denaturing isoelectric focusing showed a single band at pI 5.4. Substrate-saturation kinetics of the purified enzyme for PEP, bicarbonate, and Mg^<2+> were typical Michaelis-Menten type, with K_m-values of 60,200, and 80μM, respectively. Most effectors which are known to influence the activity of C_4- or bacterial PEPCase had only small effects on the activity of the purified enzyme at optimum pH, while some inhibitory effects by organic acids (malate, citrate and oxaloacetate) and an activating effect by glucose-6-phosphate were observed at a suboptimal pH of 7.5.
- 日本植物生理学会の論文
著者
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KOIZUMI Nozomu
Research and Education Center for Genetic Information, Nara Institute of Science and Technology
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Koizumi N
Kyoto Univ. Kyoto Jpn
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Koizumi Nozomu
Research And Education Center For Genetic Information
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Sato Fumihiko
Research Centerfor Cell and Tissue Culture, Faculty of Agriculture, Kyoto University
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Yamada Yasuyuki
Research Centerfor Cell and Tissue Culture, Faculty of Agriculture, Kyoto University
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Yamada Yukio
Fine Chemicals Research Laboratories Kaneka Corporation
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Koizumi N
Nara Inst. Sci. And Technol. Ikoma Jpn
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Koizumi Nozomu
Plant Molecular Breeding Genetics Center Nara Inst.sci.tech.
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Yamada Yukio
Mechanical Engineering Laboratory Material Science And Bioengineering Department Aist
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Sato Fumihiko
Research Center For Cell And Tissue Culture Faculty Of Agriculture Kyoto University
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Koizumi N
Laboratory Of Molecular Microbiology School Of Agriculture Nagoya University
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Yamada Yuko
Kyoto Univ. Kyoto Jpn
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Yamada Yasuyuki
Research Center For Cell And Tissue Culture Faculty Of Agriculture Kyoto University
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Sato Fumihiko
Division of Integrated Life Science, Graduate School of Biostudies, Kyoto University
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YAMADA Yasuyuki
Research Center for Ceil and Tissue Culture, Faculty of Agriculture, Kyoto University
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