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.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Research Project (R01)
Project #
3R01AA005812-18S1
Application #
6769748
Study Section
Special Emphasis Panel (ZRG4)
Program Officer
Velazquez, Jose M
Project Start
1983-04-01
Project End
2003-12-31
Budget Start
2003-07-01
Budget End
2003-12-31
Support Year
18
Fiscal Year
2003
Total Cost
$74,254
Indirect Cost
Name
Purdue University
Department
Biochemistry
Type
Schools of Earth Sciences/Natur
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907
Mukhopadhyay, Abhijit; Wei, Baoxian; Weiner, Henry (2013) Mitochondrial NAD dependent aldehyde dehydrogenase either from yeast or human replaces yeast cytoplasmic NADP dependent aldehyde dehydrogenase for the aerobic growth of yeast on ethanol. Biochim Biophys Acta 1830:3391-8
Weiner, Henry; Duester, Greg; Maser, Edmund et al. (2009) Enzymology and molecular biology of carbonyl metabolism. Introduction. Chem Biol Interact 178:1
Ho, Kwok Ki; Mukhopadhyay, Abhijit; Li, Yi Feng et al. (2008) A point mutation produced a class 3 aldehyde dehydrogenase with increased protective ability against the killing effect of cyclophosphamide. Biochem Pharmacol 76:690-6
Mukhopadhyay, Abhijit; Weiner, Henry (2007) Delivery of drugs and macromolecules to mitochondria. Adv Drug Deliv Rev 59:729-38
Mukhopadhyay, Abhijit; Yang, Chun-song; Wei, Baoxian et al. (2007) Precursor protein is readily degraded in mitochondrial matrix space if the leader is not processed by mitochondrial processing peptidase. J Biol Chem 282:37266-75
Brichac, Jiri; Ho, Kwok Ki; Honzatko, Ales et al. (2007) Enantioselective oxidation of trans-4-hydroxy-2-nonenal is aldehyde dehydrogenase isozyme and Mg2+ dependent. Chem Res Toxicol 20:887-95
Rodriguez-Zavala, Jose Salud; Allali-Hassani, Abdellah; Weiner, Henry (2006) Characterization of E. coli tetrameric aldehyde dehydrogenases with atypical properties compared to other aldehyde dehydrogenases. Protein Sci 15:1387-96
Ho, Kwok Ki; Hurley, Thomas D; Weiner, Henry (2006) Selective alteration of the rate-limiting step in cytosolic aldehyde dehydrogenase through random mutagenesis. Biochemistry 45:9445-53
Mukhopadhyay, Abhijit; Zullo, Steven J; Weiner, Henry (2006) Factors that might affect the allotopic replacement of a damaged mitochondrial DNA-encoded protein. Rejuvenation Res 9:182-90
Ho, Kwok Ki; Weiner, Henry (2005) Isolation and characterization of an aldehyde dehydrogenase encoded by the aldB gene of Escherichia coli. J Bacteriol 187:1067-73

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