Toxicological studies in yeast species : production of cytochrome P450 and evaluation of carbon tetrachloride toxicity

Atchia, Sarah Miriam (1993) Toxicological studies in yeast species : production of cytochrome P450 and evaluation of carbon tetrachloride toxicity. Doctoral thesis, University of North London.


In this work a number of yeast species were assessed for their potential in testing putative toxic and genotoxic compounds. Carbon tetrachloride (CCÍ4) was used as a model compound. Exposure of cells of Schizosaccharomyces pombe and Saccharomyces cerevisiae to carbon tetrachloride showed that the compound was toxic to the cells in a dose-dependent manner. Sch. pombe was more sensitive to the effects of carbon tetrachloride than S. -cerevisiae.

Administration of the anti-oxidant a-tocopherot (vitamin E) to ceils of Sch, pombe treated with a toxic dose of carbon tetrachloride appeared to increase cell survival. Thèse data are consistent with work carried out on mammalian cells, and indicate that the toxic effects of carbon tetrachloride are brought about via the generation of free-radical species.

Investigation of the genotoxicity of carbon tetrachloride in Sch. pombe and S. cerevisiae revealed that the compound induced mutations to chloramphenicol resistance at concentrations which were toxic to the cells. Carbon tetrachloride also induced forward mutations in Sch. pombe ade6. Genetic analysis of chloramphenicol resistant mutants indicated non-Mendelian inheritance.

The drug-metabolising enzyme cytochrome P450 is thought to metabolise carbon tetrachloride to toxic species. The ability of carbon tetrachloride to act as a substrate for the haemoprotein was indicated by the type 1 binding spectrum produced in Sch. pombe. a-Tocopherol was found to increase levels of cytochrome P450. The production of cytochrome P450 was investigated in the three yeast species Sch. pombe, S. cerevisiae and Candida parapsitosis. The enzyme was optimally produced under conditions of glucose repression during the late logarithmic phase of growth. Levels of the enzyme were consistently found to be greater in respiratory deficient (petite) mutants of the yeast S. cerevisiae than in wild type strains.

The enzyme was isolated from microsomal preparations of C. parapsilosis. SDS-PAGE analysis revealed a major band in the solubilised fraction possessing a molecular mass of approximately 48 kDa which was tentatively identified as cytochrome P450.

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