盐生植物盐地碱蓬酪氨酸酶的酶学特性

酪氨酸酶是植物甜菜素生物合成的限速酶,但是,其酶学特性尚不了解。以黑暗培养3d的盐地碱蓬幼苗为材料,采用NaF抽提、饱和硫酸铵沉淀法提取盐地碱蓬中的酪氨酸酶,以研究其酶学特性。结果表明,酪氨酸酶氧化活性的最适温度为35℃,最适pH值为6.6,最适条件下Km=1.09mmol·L-1,Vmax=71.43μmol·g-1（FW）·min-1;酪氨酸酶羟化活性的最适温度为40℃,最适pH值为6.6,最适条件下Km=3.16mmol·L-1,Vmax=0.645μmol·g-1（FW）·min-1。Na2S2O3是盐地碱蓬酪氨酸酶的强效抑制剂,0.05mol·L-1Na2S2O3几乎完全抑制酪氨酸酶氧化及羟化活性。而0.01mmol·L-1的Cu2＋可以显著激活酪氨酸酶的氧化及羟化活性,分别为对照的126%和128.2%。这些结果表明盐地碱蓬中酪氨酸酶的羟化活性是影响甜菜红素合成速率的关键,也为深入研究盐地碱蓬酪氨酸酶在甜菜素合成中的作用及其与环境之间的关系奠定了基础。

Enzymatic Characteristics of Tyrosinase in Euhalophyte Suaeda salsa

Tyrosinase is the first key enzyme in betalains biosynthesis of plants while its enzymatic characteristics remain unclear.In order to investigate the properties of tyrosinase,3-day-old Suaeda salsa seedlings（cultured in dark） were used and the tyrosinase was partially purified by ammonium sulphate （NH4）2SO4] precipitation after being extracted by sodium fluoride（NaF）.The results indicated that,in terms of O-diphenol oxidation activity,the optimum temperature and pH values were 35 ℃ and 6.6,the Km=1.09 mmol·L-1 and Vmax=71.43 μmol·g-1（FW）·min-1;and as for the activity of monophenol hydroxylation,the optimum temperature and pH value were 40 ℃ and 6.6,the Km=3.16 mmol·L-1 and Vmax=0.645 μmol·g-1（FW）·min-1.Na2S2O3 was the potent inhibitor of tyrosinase,and 0.05 mol·L-1 Na2S2O3 almost completely inhibited the activity of O-diphenol oxidation and monophenol hydroxylation.Moreover,the activity of O-diphenol oxidation and monophenol hydroxylation was significantly activated by 0.01 mmol·L-1 Cu2＋,126%、128.2% of control respectively.These results suggested that tyrosinase hydroxylation activity of S.salsa played a key role in betalains biosynthesis and laid the foundation of deep investigating the role of tyrosinase in betalains biosynthesis in plants and its interaction with environment factors.