Aromatic metabolism in plants III. Quinate dehydrogenase from mung bean cell suspension cultures

Oluf L. Gamborg
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引用次数: 25

Abstract

Quinate dehydrogenase (Quinate: NAD+ oxidoreductase, EC 1.1.1.24) was extracted from liquid suspension cultures of mung bean (Phaseolus aureus Roxb.).

The cells were disrupted by ultrasonic energy, and the enzyme purified by precipitation with ammonium sulfate and elution from columns of Sephadex G-50 and hydroxylapatite. The enzyme was unstable but the activity could be maintained for several weeks after purification of hydroxylapatite when stored at pH 7.5 and at −20°.

The optimum pH for activity was 9.6. The enzyme was specific for NAD+. Addition of phenylpyruvate, phenylalanine, cinnamate, or shikimate had no effect on its activity. The enzyme was inhibited by sulfhydryl inhibitors, borate, molybdate, and dehydroquinate. 4-Hydroxybenzoic acid and 3-hydroxybenzoic acid were competitive inhibitors.

The cells also contained 5-dehydroquinate dehydratase (EC 4.2.1.10) and shikimate dehydrogenase (EC 1.1.1.25) and thus contained all the enzymes necessary for the interconversion of quinate and shikimate. Quinate dehydrogenase resembled shikimate dehydrogenase in its pH optimum, inhibition by borate and by the substituted benzoic acids.

植物的芳香代谢3。绿豆细胞悬浮培养的奎宁脱氢酶
从绿豆(Phaseolus aureus Roxb.)液体悬浮培养液中提取Quinate脱氢酶(Quinate: NAD+ oxidoreductase, EC 1.1.1.24)。用超声能量破坏细胞,用硫酸铵沉淀和从Sephadex G-50和羟基磷灰石柱中洗脱纯化酶。羟基磷灰石纯化后,在pH 7.5和- 20°条件下,酶活性可维持数周。最适pH为9.6。这种酶对NAD+具有特异性。添加苯丙酮酸、苯丙氨酸、肉桂酸或莽草酸对其活性无影响。巯基抑制剂、硼酸盐、钼酸盐和脱氢奎酸盐均能抑制该酶。4-羟基苯甲酸和3-羟基苯甲酸是竞争性抑制剂。该细胞还含有5-脱氢quinate脱氢酶(EC 4.2.1.10)和莽草酸脱氢酶(EC 1.1.1.25),因此包含了所有需要的酶的相互转化的quinate和莽草酸。喹酸脱氢酶与莽草酸脱氢酶的最佳pH值相似,硼酸盐和取代苯甲酸对其有抑制作用。
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