Translator Disclaimer
1 August 2005 Identification of Cytochrome-b5 Reductase as the Enzyme Responsible for NADH-Dependent Lucigenin Chemiluminescence in Human Spermatozoa
Author Affiliations +
Abstract

Lucigenin-dependent chemiluminescence together with 2-[4-iodophenyl]-3-[4-nitrophenyl]-5-[2,4-disulfophenyl]-2H tetrazolium monosodium salt (WST-1) reduction can be detected following addition of NADH to many cell types, including human sperm suspensions. Although many reports suggest that such a phenomenon is due to reactive oxygen species production, other oxygen detecting metabolite probes, such as MCLA and luminol, do not produce a chemiluminescent signal in this model system. The enzyme responsible for NADH-dependent lucigenin chemiluminescence was purified and identified as cytochrome-b5 reductase. In support of this concept, COS-7 cells overexpressing cytochrome-b5 reductase displayed at least a 3-fold increase in the previously mentioned activity compared with mock-transfected cells. Fractions containing cytochrome-b5 reductase were capable of inducing both lucigenin-dependent chemiluminescence and WST-1 reduction. Oxygen radicals clearly did not mediate the cytochrome b5-mediated activation of these probes in vitro since neither luminol nor MCLA gave a chemiluminescence response in the presence of the enzyme and the cofactor NADH. These results emphasize the importance of the direct NADH-dependent reduction of these putative superoxide-sensitive probes by cytochrome-b5 reductase even though this enzyme does not, on its own accord, produce reactive oxygen species.

Mark A. Baker, Anton Krutskikh, Benjamin J. Curry, Louise Hetherington, and R. John Aitken "Identification of Cytochrome-b5 Reductase as the Enzyme Responsible for NADH-Dependent Lucigenin Chemiluminescence in Human Spermatozoa," Biology of Reproduction 73(2), 334-342, (1 August 2005). https://doi.org/10.1095/biolreprod.104.037960
Received: 10 November 2004; Accepted: 1 April 2005; Published: 1 August 2005
JOURNAL ARTICLE
9 PAGES

This article is only available to subscribers.
It is not available for individual sale.
+ SAVE TO MY LIBRARY

SHARE
ARTICLE IMPACT
RIGHTS & PERMISSIONS
Get copyright permission
Back to Top