关于5'-磷酸吡哆醛结合时天冬氨酸氨基转移酶的蛋白质荧光和酶活性变化的研究。
Studies on the changes in protein fluorescence and enzymic activity of aspartate aminotransferase on binding of pyridoxal 5'-phosphate.
作者信息
Evans R W, Holbrook J J
出版信息
Biochem J. 1974 Dec;143(3):643-9. doi: 10.1042/bj1430643.
Abstract
- The alpha and beta subforms of aspartate aminotransferase were purified from pig heart. 2. The alpha subform contained 2mol of pyridoxal 5'-phosphate. The apo-(alpha subform) could be fully reactived by combination with 2mol of cofactor. 3. The protein fluorescence of the apo-(alpha subform) decreased non-linearly with increase in enzyme activity and concentration of bound cofactor. 4. It is concluded that the enzyme activity/mol of bound cofactor is largely independent of the number of cofactors bound to the dimer. 5. The beta subform had approximately half the specific enzyme activity of the alpha subform, and contained an average of one active pyridoxal 5'-phosphate molecule per molecule, which could be removed by glutamate, and another inactive cofactor which could only be removed with NaOH. 6. On recombination with pyridoxal 5'-phosphate the protein fluorescence of the apo-(beta subform) decreased linearly, showing that each dimeric enzyme molecule contained one active and one inactive bound cofactor. 7. The results are not consistent with a flip-flop mechanism for this enzyme.
摘要
- 从猪心中纯化出天冬氨酸转氨酶的α和β亚基形式。2. α亚基含有2摩尔的磷酸吡哆醛5′-磷酸。脱辅基（α亚基）可通过与2摩尔辅因子结合而完全重新激活。3. 脱辅基（α亚基）的蛋白质荧光随酶活性和结合辅因子浓度的增加呈非线性下降。4. 得出的结论是，每摩尔结合辅因子的酶活性在很大程度上与结合到二聚体上的辅因子数量无关。5. β亚基的比酶活性约为α亚基的一半，每个分子平均含有一个活性磷酸吡哆醛5′-磷酸分子，可被谷氨酸去除，还有另一个无活性的辅因子，只能用氢氧化钠去除。6. 脱辅基（β亚基）与磷酸吡哆醛5′-磷酸重新结合时，其蛋白质荧光呈线性下降，表明每个二聚体酶分子含有一个活性和一个无活性的结合辅因子。7. 这些结果与该酶的翻转机制不一致。
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