吡哆醛激酶（Pyridoxal Kinase, PLK）是维生素B6（VB6）代谢关键酶。从茶鲜叶中直接提取含PLK的粗酶液,经过硫酸铵沉淀、透析处理后,采用苯肼衍生化方法检测定PLK活性,并对其基本酶学性质进行分析。结果表明,提取时加入100%茶鲜重的PVPP,效果最佳。纯化后的茶鲜叶PLK比活力为0.737 nmol/(min·mg),纯化倍数为4.0;当有二价金属离子存在时,PLK才具有催化活性,其中Zn2+对其催化作用最强。该酶的最适反应温度为60℃;在pH6.0左右酶活力最高。在最适反应条件下,测得反应底物ATP和PL的Km值分别为19.3 μmol/L和53 μmol/L。
Pyridoxal Kinase (PLK) is the key enzyme in VB6 metabolism. The crude solution of PLK enzyme was directly extracted from tea plant, after depositing and dialysing with ammonia sulfate by using the phenylhydrazine derivative method to assay the activity of the PLK and its properties. Results showed that the addition of 100%(m/m) PVPP of the fresh tea leaves extraction the PLK could get the best results. The purified tea PLK was enriched by 4.0 fold and its specific activity was 0.737 nmol/(min·mg protein). The requirement of divalent ion for catalysis was mandatory, Zn2+ was the most efficient ion for catalysis; the optimum temperature and pH of the enzyme were 60℃ and 6.0, respectively. Under optimal conditions, the Km values for pyridoxal and its reactive substrate ATP were 19.3 μmol/L and 53 μmol/L.
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