Hall L R, Hanzlik R P
Department of Medicinal Chemistry, University of Kansas, Lawrence 66045-2506.
J Biol Chem. 1990 Jul 25;265(21):12349-55.
Liver microsomal cytochrome P-450 readily N-dealkylates N,N-dimethylamides. N-Methyl-N-hydroxymethyl amides were isolated as intermediates and characterized by gas chromatography-mass spectrometry as their trimethylsilyl ethers. Intramolecular kinetic deuterium isotope effects measured for the enzymic N-demethylation of a series of 12 aromatic and aliphatic N-methyl-N-trideuteriomethyl amides, RCON(CH3)CD3, varied from 3.6 to 6.9 but were independent of both amide bond rotation rate and substrate oxidation potential. These values, which represent a lower limit to the intrinsic isotope effect (Dkintrinsic), are significantly larger than those observed for anodic N-demethylation and are consistent with a mechanism involving hydrogen atom abstraction. On the other hand, with N,N-dimethylbenzamide the intermolecular kinetic deuterium isotope effects on Vmax and Vmax/Km were found to be much smaller (1.23 and 1.75, respectively) indicating substantial suppression of the intrinsic isotope effect. Such suppression indicates the occurrence of a rate-limiting step other than the isotopically sensitive step together with a strong commitment to catalysis.
肝脏微粒体细胞色素P - 450能轻易地使N，N - 二甲基酰胺发生N - 脱烷基反应。N - 甲基 - N - 羟甲基酰胺作为中间体被分离出来，并通过气相色谱 - 质谱法将其表征为三甲基硅醚。对一系列12种芳香族和脂肪族N - 甲基 - N - 三氘代甲基酰胺RCON(CH3)CD3进行酶促N - 脱甲基反应的分子内动力学氘同位素效应测定，其值在3.6至6.9之间变化，但与酰胺键旋转速率和底物氧化电位均无关。这些值代表了固有同位素效应（Dkintrinsic）的下限，明显大于阳极N - 脱甲基反应所观察到的值，并且与涉及氢原子提取的机制一致。另一方面，对于N，N - 二甲基苯甲酰胺，发现对Vmax和Vmax/Km的分子间动力学氘同位素效应要小得多（分别为1.23和1.75），这表明固有同位素效应受到了显著抑制。这种抑制表明除了同位素敏感步骤之外还存在一个限速步骤，以及对催化作用的强烈倾向。
