The hexose monophosphate (or pentose phosphate) pathway and cytosolic isocitrate dehydrogenase (IDP2) have been found to be essential adjuncts to catalase function in preventing intracellular accumulation of the hydrogen peroxide generated during beta-oxidation of fatty acids in peroxisomes. The absence of these two cytosolic sources of NADPH results in lethality of yeast cells transferred to media with fatty acid carbon sources. These observations, peroxisomal metabolism as a source of lethal levels of hydrogen peroxide and a requirement for cytosolic NADPH to prevent lethality, are novel observations in the area of cellular oxidative stress. Further investigation of these phenomena is proposed because oxidative damage to cellular macromolecules has been clearly associated with degenerative diseases and with the process of aging.
Aims of this proposal are: (A) Similar requirements for cytosolic NADPH in removal of oxidative byproducts generated during mitochondrial respiration will be examined using a collection of Saccharomyces cerevisiae mutants with combinations of disruptions in genes encoding these and other enzymes with specific antioxidant functions. (B) The heritability of damage to specific genes and cellular proteins will be assessed using genetic crosses of yeast mutant strains that exhibit similar growth phenotypes as a result of endogenous oxidative damage. (C) The unique antioxidant functions of IDP2 will be explored by replacement with homologous yeast isozymes and by using yeast two-hybrid assays to test and screen for specific interactions with other cellular proteins. (D) Patterns of expression and cellular localization of mammalian IDP2 will be examined to clarify possible roles in support of peroxisomal beta- oxidation and/or in provision of NADPH for antioxidant functions.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Project (R01)
Project #
5R01AG017477-04
Application #
6629856
Study Section
Medical Biochemistry Study Section (MEDB)
Program Officer
Finkelstein, David B
Project Start
2000-03-01
Project End
2005-02-28
Budget Start
2003-03-15
Budget End
2005-02-28
Support Year
4
Fiscal Year
2003
Total Cost
$247,120
Indirect Cost
Name
University of Texas Health Science Center San Antonio
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
800772162
City
San Antonio
State
TX
Country
United States
Zip Code
78229
Lu, Qian; McAlister-Henn, Lee (2010) Peroxisomal localization and function of NADP+ -specific isocitrate dehydrogenases in yeast. Arch Biochem Biophys 493:125-34
Minard, Karyl I; McAlister-Henn, L (2009) Redox responses in yeast to acetate as the carbon source. Arch Biochem Biophys 483:136-43
Garcia, Joshua A; Minard, Karyl I; Lin, An-Ping et al. (2009) Disulfide bond formation in yeast NAD+-specific isocitrate dehydrogenase. Biochemistry 48:8869-78
Lu, Qian; Minard, Karyl I; McAlister-Henn, Lee (2008) Dual compartmental localization and function of mammalian NADP+-specific isocitrate dehydrogenase in yeast. Arch Biochem Biophys 472:17-25
Lin, An-Ping; Hakala, Kevin W; Weintraub, Susan T et al. (2008) Suppression of metabolic defects of yeast isocitrate dehydrogenase and aconitase mutants by loss of citrate synthase. Arch Biochem Biophys 474:205-12
Minard, Karyl I; Carroll, Christopher A; Weintraub, Susan T et al. (2007) Changes in disulfide bond content of proteins in a yeast strain lacking major sources of NADPH. Free Radic Biol Med 42:106-17
Minard, Karyl I; McAlister-Henn, Lee (2005) Sources of NADPH in yeast vary with carbon source. J Biol Chem 280:39890-6
Contreras-Shannon, Veronica; Lin, An-Ping; McCammon, Mark T et al. (2005) Kinetic properties and metabolic contributions of yeast mitochondrial and cytosolic NADP+-specific isocitrate dehydrogenases. J Biol Chem 280:4469-75
Contreras-Shannon, Veronica; McAlister-Henn, Lee (2004) Influence of compartmental localization on the function of yeast NADP+-specific isocitrate dehydrogenases. Arch Biochem Biophys 423:235-46
McCammon, Mark T; McAlister-Henn, Lee (2003) Multiple cellular consequences of isocitrate dehydrogenase isozyme dysfunction. Arch Biochem Biophys 419:222-33

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