Blytt H J, Brotherton J E, Butler L
Anal Biochem. 1985 Jun;147(2):517-20. doi: 10.1016/0003-2697(85)90307-0.
We report a new procedure for isolating a covalent phosphoryl enzyme (diester) intermediate of bovine intestinal 5'-nucleotide phosphodiesterase. The convenience of the procedure makes it possible to determine effects of reaction conditions on the yield of covalent intermediate. Under optimum conditions, using [methyl-3H]deoxythymidine 5'-triphosphate as substrate, more than 50% of the enzyme is recovered as thymidylyl enzyme, a 10-fold increase in yield over the previous procedure (M. Landt and L. G. Butler, 1978, Biochemistry 17, 4130-4135). Yields of thymidylyl enzyme were maximal at pH 4, whereas optimum catalytic activity is observed at pH greater than 9.
我们报道了一种分离牛肠5'-核苷酸磷酸二酯酶共价磷酰化酶（二酯）中间体的新方法。该方法的便利性使得确定反应条件对共价中间体产量的影响成为可能。在最佳条件下，以[甲基-3H]脱氧胸苷5'-三磷酸为底物，超过50%的酶以胸苷酰化酶形式回收，产量比之前的方法提高了10倍（M. Landt和L. G. Butler，1978年，《生物化学》17，4130 - 4135）。胸苷酰化酶的产量在pH 4时最高，而最佳催化活性在pH大于9时观察到。
