Isolation, nucleotide sequence, and disruption of the Saccharomyces cerevisiae gene encoding mitochondrial NADP(H)-specific isocitrate dehydrogenase

Robert J. Haselbeck, Lee McAlister-Henn

Research output: Contribution to journalArticle

60 Citations (Scopus)

Abstract

Mitochondrial NADP(H)-specific isocitrate dehydrogenase (IDP1) was purified from yeast cells grown with acetate as a carbon source. IDP1 was shown to be a dimer with a subunit molecular weight of approximately 45,000. Immunochemical levels of IDP1 were found to vary in inverse proportion with those of mitochondrial NAD(H)-specific isocitrate dehydrogenase in cells grown with glucose or with acetate as a carbon source. A 20-residue amino-terminal sequence was obtained for IDP1, and degenerate oligonucleotides were used to synthesize a 50-base pair polymerase chain reaction product corresponding to the coding region for a portion of the amino terminus. The 50-base pair DNA fragment was used as a hybridization probe to identify plasmids containing the IDP1 gene in a yeast genomic DNA library. The complete nucleotide sequence of the IDP1 coding region was determined and translated into a 412-residue amino acid sequence for the mature protein which is preceded by a putative 16-residue mitochondrial targeting presequence. A haploid yeast strain containing a chromosomal disruption of the IDP1 locus was constructed and found to be capable of growth with glucose but not with other carbon sources, suggesting that IDP1 provides a critical function and may be the primary source of NADPH in yeast mitochondria.

Original languageEnglish (US)
Pages (from-to)2339-2345
Number of pages7
JournalJournal of Biological Chemistry
Volume266
Issue number4
StatePublished - Feb 5 1991
Externally publishedYes

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Isocitrate Dehydrogenase
Gene encoding
Mitochondrial Genes
NADP
Yeast
Saccharomyces cerevisiae
Nucleotides
Yeasts
Carbon
Base Pairing
Acetates
Cells
Glucose
Mitochondria
Genomic Library
Haploidy
Polymerase chain reaction
Reaction products
Gene Library
Oligonucleotides

ASJC Scopus subject areas

  • Biochemistry

Cite this

Isolation, nucleotide sequence, and disruption of the Saccharomyces cerevisiae gene encoding mitochondrial NADP(H)-specific isocitrate dehydrogenase. / Haselbeck, Robert J.; McAlister-Henn, Lee.

In: Journal of Biological Chemistry, Vol. 266, No. 4, 05.02.1991, p. 2339-2345.

Research output: Contribution to journalArticle

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