Faculty Collaboration Database

Enzymatic reducibility in relation to cytotoxicity for various cholesterol hydroperoxides. Biochemistry 1996 Jul 02;35(26):8670-9

Date

07/02/1996

Pubmed ID

8679629

DOI

10.1021/bi960522k

Scopus ID

2-s2.0-0029945390 (requires institutional sign-in at Scopus site)   43 Citations

Abstract

Phospholipid hydroperoxide glutathione peroxidase (PHGPX) is a selenoenzyme that can catalyze the direct reduction of various membrane lipid hydroperoxides and by so doing could play a vital role in cytoprotection against peroxidative damage. The activity of purified testicular PHGPX on several photochemically-generated cholesterol hydroperoxide (ChOOH) species was investigated, using high-performance liquid chromatography with electrochemical detection for peroxide analysis and thinlayer chromatography with 14C-radiodetection for diol product analysis. The following ChOOH isomers were monitored: 5 alpha-OOH, 6 alpha-OOH, 6 beta-OOH (singlet oxygen adducts), and unresolved 7 alpha,7 beta-OOH (derived from 5 alpha-OOH rearrangement). Apparent first-order rate constants for GSH/PHGPX-induced peroxide loss (or diol accumulation) in Triton X-100 micelles, unilamellar liposomes, or erythrocyte ghost membranes increased in the following order: 5 alpha-OOH < 6 alpha-OOH approximately equal to 7 alpha,7 beta-OOH < 6beta-OOH. A similar trend was observed when the peroxides were incubated with Triton Iysates of Se-replete L1210 or K562 cells, implicating PHGPX in these reactions. Consistent with this, there was little or no ChOOH reduction if GSH was omitted or if lysates from Se-deprived cells were used. Liposomal 5 alpha-OOH was found to be much more cytotoxic than equimolar liposomal 6 beta-OOH, producing a 50% loss of L1210 clonogenicity at approximately 1/5 the concentration of the latter. Faster uptake of 5 alpha-OOH was ruled out as the basis for greater cytotoxicity, suggesting that relatively inefficient metabolism by the GSH/PHGPX system might be the reason. As supporting evidence, it was found that cells accumulate the diol reduction product of 5 alpha-OOH more slowly than that of 6 beta-OOH during incubation with the respective peroxides. Slow detoxification coupled with rapid formation makes 5 alpha-OOH potentially the most damaging ChOOH to arise in cells exposed to singlet oxygen.

Author List

Korytowski W, Geiger PG, Girotti AW

Author

Albert W. Girotti PhD Adjunct Professor in the Biochemistry department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Catalysis
Cell Line
Cell Survival
Cholesterol
Glutathione Peroxidase
Humans
Kinetics
Mice
Oxidation-Reduction
Oxygen
Singlet Oxygen