二氢鱼藤酮与电子传递链复合体I结合的定量放射自显影术。

Quantitative autoradiography of dihydrorotenone binding to complex I of the electron transport chain.

作者信息

Greenamyre J T, Higgins D S, Eller R V

机构信息

Department of Neurology, University of Rochester Medical Center, New York 14642.

出版信息

J Neurochem. 1992 Aug;59(2):746-9. doi: 10.1111/j.1471-4159.1992.tb09431.x.

DOI:10.1111/j.1471-4159.1992.tb09431.x

PMID:1629744

Abstract

Defective complex I activity has been linked to Parkinson's disease and Huntington's disease, but little is known of the regional distribution of this enzyme in the brain. We have developed a quantitative autoradiographic assay using [3H]dihydrorotenone ([3H]DHR) to label and localize complex I in brain tissue sections. Binding was specific and saturable and in the cerebellar molecular layer had a KD of 11.5 +/- 1.3 nM and a Bmax of 11.0 +/- 0.4 nCi/mg of tissue. Unlabeled rotenone and 1-methyl-4-phenylpyridinium ion competed effectively for DHR binding sites. Binding was markedly enhanced by 100 microM NADH. The distribution of complex I in brain, as revealed by DHR autoradiography, is unique but somewhat similar to that of cytochrome oxidase (complex IV). This assay may provide new insight into the roles of complex I in brain function and neurodegeneration.

摘要

复合体I活性缺陷与帕金森病和亨廷顿病有关，但对于该酶在大脑中的区域分布了解甚少。我们开发了一种定量放射自显影测定法，使用[3H]二氢鱼藤酮（[3H]DHR）标记并定位脑组织切片中的复合体I。结合是特异性的且具有饱和性，在小脑分子层中，KD为11.5±1.3 nM，Bmax为11.0±0.4 nCi/mg组织。未标记的鱼藤酮和1-甲基-4-苯基吡啶离子可有效竞争DHR结合位点。100μM NADH可显著增强结合。DHR放射自显影显示的复合体I在大脑中的分布独特，但与细胞色素氧化酶（复合体IV）的分布有些相似。该测定法可能为复合体I在脑功能和神经退行性变中的作用提供新的见解。