Effective NADH-dependent oxidation of 7β-hydroxy-Δ8-tetrahydrocannabinol to the corresponding ketone by Japanese monkey hepatic microsomes
Effective NADH-dependent oxidation of 7β-hydroxy-Δ8-tetrahydrocannabinol to the corresponding ketone by Japanese monkey hepatic microsomes
复制标题
DOI：
10.1248/bpb.28.646
复制
发表时间：
2005-04-01
影响因子：
2
通讯作者：
Yamamoto, I
中科院分区：
文献类型：
--
作者：
Matsunaga, T;Higuchi, S;Yamamoto, I
日本猴肝微粒体从7 β-羟基-δ（8）-THC生成7-氧代-δ（8）-四氢大麻酚（7-氧代-δ（8）-THC）的依赖于NADPH的活性，在底物浓度为72.7 μ m时，表现出约70%的依赖于NADPH的活性（100%），尽管NADPH是最大活性的必需辅因子。典型的P450抑制剂如SKF 525-A和甲吡酮均能显著抑制NADH和NADPH依赖的活性。这两种活性几乎完全抑制NADPH-P450还原酶抑制剂氯化二苯基碘。根据底物浓度的比例的NADH和NADPH依赖的活动变化显着。有趣的是，在13-50 μ M的低底物浓度下，NADH依赖的活性高于NADPH。该比率还受辅因子浓度的影响。在从日本猴肝微粒体中纯化的CYP 3A 8作为负责NADPH依赖性氧化的主要酶的重构系统中，NADH和NADPH可以维持7 β-羟基-δ（8）-THC氧化为相应的酮。猴肝对7 β-羟基-δ（8）-THC的NADH依赖性氧化主要由CYP 3A 8催化，NADPH依赖性氧化也是如此。这些结果表明，作为辅因子的NADH也可用于7 β-羟基-δ（8）-THC的氧化，并且反应的辅因子要求随底物和/或辅因子的浓度而变化。
The NADH-dependent activity by hepatic microsomes of Japanese monkeys for 7-oxo-Delta(8)-tetrahydro-cannabinol (7-oxo-Delta(8)-THC) formation from 7 beta-hydroxy-Delta(8)-THC exhibited about 70% of the NADPH-dependent activity (100%) at the substrate concentration of 72.7 mu m, although NADPH was an obligatory cofactor for maximal activity. Both NADH- and NADPH-dependent activities were significantly inhibited by the typical P450 inhibitors, such as SKF525-A and metyrapone. Both activities were almost completely inhibited by the NADPH-P450 reductase inhibitor diphenyliodonium chloride. The ratio of NADH- and NADPH-dependent activities varied significantly according to the substrate concentration. Interestingly, the NADH-dependent activity was higher than that of NADPH at low substrate concentrations of 13-50 mu m. The ratio was also affected by the cofactor concentration. In the reconstituted system of CYP3A8 purified from hepatic microsomes of Japanese monkeys as a major enzyme responsible for the NADPH-dependent oxidation, NADH as well as NADPH could sustain the oxidation of 7 beta-hydroxy-Delta(8)-THC to the corresponding ketone. The NADH-dependent oxidation of 7 beta-hydroxy-Delta(8)-THC by monkey livers is mainly catalyzed by CYP3A8 as well as the NADPH-dependent oxidation. These results indicate that NADH as a cofactor may be also useful for the oxidation of 7 beta-hydroxy-Delta(8)-THC, and that the cofactor requirement for the reaction is varied by the concentrations of substrate and/or cofactor.