Lehman T C, Hale D E, Bhala A, Thorpe C
Department of Chemistry, University of Delaware, Newark 19716.
Anal Biochem. 1990 May 1;186(2):280-4. doi: 10.1016/0003-2697(90)90080-s.
A sensitive assay for medium chain acyl-CoA dehydrogenase has been developed by substituting ferricenium hexafluorophosphate for the physiological acceptor, electron transferring flavoprotein. The ferricenium ion is a facile oxidant of the octanoyl-CoA-reduced enzyme with a Vmax of 1400 min-1 and a KM of 55 microM at pH 7.6. The ferricenium assay does not require additional mediator dyes, exhibits low background rates, and avoids the necessity of purifying substantial amounts of electron transferring flavoprotein. Unlike the fluorescence-based electron transferring flavoprotein assay, this new procedure can be performed aerobically. Both assays give comparable results when tested with crude fibroblast homogenates from normal and medium chain acyl-CoA dehydrogenase deficient patients. The convenience of the ferricenium method suggests it may be generally useful as a screening assay for a number of acyl-CoA dehydrogenases.
通过用六氟磷酸铁鎓替代生理受体电子传递黄素蛋白，已开发出一种灵敏的中链酰基辅酶A脱氢酶检测方法。在pH 7.6条件下，铁鎓离子是辛酰辅酶A还原酶的一种简便氧化剂，Vmax为1400 min-1，KM为55 μM。铁鎓检测法不需要额外的介体染料，背景速率低，且无需纯化大量的电子传递黄素蛋白。与基于荧光的电子传递黄素蛋白检测法不同，这种新方法可以在有氧条件下进行。用来自正常和中链酰基辅酶A脱氢酶缺乏患者的成纤维细胞粗匀浆进行测试时，两种检测方法给出的结果相当。铁鎓方法的便利性表明它可能普遍适用于多种酰基辅酶A脱氢酶的筛选检测。
