ENZYMOLOGY/MOLECULAR BIOLOGY OF ALDEHYDE DEHYDROGENASE

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DESCRIPTION:(Adapted from the Investigator's Abstract) Aldehyde dehydrogenases are generally detoxifying enzymes necessary to remove toxic aldehydes from the cell. The aldehydes range from acetaldehyde formed during the oxidation of ingested alcoholic beverages, to some formed from the action of light on poly unsaturated fats. Recently it was found that some drugs used in chemotherapy form a cytotoxic aldehyde and will kill cells void of the enzymes, such as some tumor cells and bone marrow cells. Yeast contains its own aldehyde dehydrogenases while HeLa cells do not have the enzyme. The investigators will delete the enzyme from yeast so these cells can be transformed with the human liver forms of the enzyme and be used to make mutations in them. These will be used for mechanistic studies and to try to develop an enzyme more capable of protecting a cell against the toxic effects of drug such as cyclophophamides. HeLa cells will be changed so that they can metabolize ethanol. These cells will be transformed with modified aldehyde dehydrogenases so they can be used to study the metabolism of acetaldehyde in the presence of mutated enzymes they will produce. Many Oriental people suffer reactions when they consume alcoholic beverages. These people possess an altered, inactive form of the mitochondrial enzyme. However, they also possess copies of the active form of the enzyme. A new method will be proposed to produce these heterotetramers of the enzyme so the investigators can study how an inactive subunit affects the activity of the active subunit. The cells will also be used to determine in vivo ability of the mutated enzymes to protect them. A mutational approach will be employed so the investigators can study questions related to the mechanism of action of the enzyme, with the long term goal of being able to design aldehyde dehydrogenases which might possess more desirable properties when used in gene replacement therapy or to make transgenic animals.