Blackwell L F, Motion R L, MacGibbon A K, Hardman M J, Buckley P D
Biochem J. 1987 Mar 15;242(3):803-8. doi: 10.1042/bj2420803.
The displacement of NADH from the aldehyde dehydrogenase X NADH complex by NAD+ was followed at pH 7.0, and the data were fitted by a non-linear least-squares iterative procedure. At pH 7.0 the decay constants for the dissociation of NADH from aldehyde dehydrogenase X NADH complexes (1.62 +/- 0.09 s-1 and 0.25 +/- 0.004 s-1) were similar to the values previously determined by MacGibbon, Buckley & Blackwell [(1977) Biochem. J. 165, 455-462] at pH 7.6, and apparent differences between these values and those reported by Dickinson [(1985) Biochem. J. 225, 159-165] are resolved. Experiments at low concentrations of propionaldehyde show that isomerization of a binary E X NADH complex is part of the normal catalytic mechanism of the enzyme. Evidence is presented that the active-site concentration of aldehyde dehydrogenase is halved when enzyme is pre-diluted to low concentrations before addition of NAD+ and substrate. The consequences of this for the reported values of kcat. are discussed. A general mechanism for the aldehyde dehydrogenase-catalysed oxidation of propionaldehyde which accounts for the published kinetic data, at concentrations of aldehyde which bind only at the active site, is presented.
在pH 7.0条件下，研究了NAD⁺从醛脱氢酶X NADH复合物中置换NADH的过程，并通过非线性最小二乘法迭代程序对数据进行拟合。在pH 7.0时，NADH从醛脱氢酶X NADH复合物解离的衰减常数（1.62±0.09 s⁻¹和0.25±0.004 s⁻¹）与MacGibbon、Buckley和Blackwell [（1977年）《生物化学杂志》165卷，455 - 462页] 在pH 7.6时先前测定的值相似，这些值与Dickinson [（1985年）《生物化学杂志》225卷，159 - 165页] 报道的值之间的明显差异得到了解决。低浓度丙醛的实验表明，二元E X NADH复合物的异构化是该酶正常催化机制的一部分。有证据表明，在加入NAD⁺和底物之前将酶预稀释至低浓度时，醛脱氢酶的活性位点浓度减半。讨论了这对报道的kcat值的影响。提出了一种醛脱氢酶催化丙醛氧化的通用机制，该机制解释了在仅在活性位点结合的醛浓度下已发表的动力学数据。
