Abstract
When baker's yeast spheroplasts were lysed by mild osmotic shock, practically all of the isopropylmalate isomerase and the β-isopropylmalate dehydrogenase was released into the 30,000 × g supernatant fraction, as was the cytosol marker enzyme, glucose-6-phosphate dehydrogenase. α-Isopropylmalate synthase, however, was not detected in the initial supernatant, but could be progressively solubilized by homogenization, appearing more slowly than citrate synthase but faster than cytochrome oxidase. Of the total glutamate-α-ketoisocaproate transaminase activity, approximately 20% was in the initial soluble fraction, whereas solubilization of the remainder again required homogenization of the spheroplast lysate. Results from sucrose density gradient centrifugation of a cell-free particulate fraction and comparison with marker enzymes suggested that α-isopropylmalate synthase was located in the mitochondria. It thus appears that, in yeast, the first specific enzyme in the leucine biosynthetic pathway (α-isopropylmalate synthase) is particulate, whereas the next two enzymes in the pathway (isopropylmalate isomerase and β-isopropylmalate dehydrogenase) are “soluble,” with glutamate-α-ketoisocaproate transaminase activity being located in both the cytosol and particulate cell fractions.
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- Altmiller D. H. Neurospora mutants with mitochondria deficient in dihydroxy acid dehydratase. Properties of dihydroxy acid dehydratase from mutant strain 332. Biochem Biophys Res Commun. 1972 Nov 15;49(4):1000–1006. doi: 10.1016/0006-291x(72)90311-7. [DOI] [PubMed] [Google Scholar]
- Bernhardt S. A., Davis R. H. Carbamoyl phosphate compartmentation in Neurospora: histochemical localization of aspartate and ornithine transcarbamoylases. Proc Natl Acad Sci U S A. 1972 Jul;69(7):1868–1872. doi: 10.1073/pnas.69.7.1868. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cassady W. E., Leiter E. H., Bergquist A., Wagner R. P. Separation of mitochondrial membranes of Neurospora crassa. II. Submitochondrial localization of the isoleucine-valine biosynthetic pathway. J Cell Biol. 1972 Apr;53(1):66–72. doi: 10.1083/jcb.53.1.66. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Collier R. H., Kohlhaw G. Nonidentity of the aspartate and the aromatic aminotransferase components of transaminase A in Escherichia coli. J Bacteriol. 1972 Oct;112(1):365–371. doi: 10.1128/jb.112.1.365-371.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- DUELL E. A., INOUE S., UTTER M. F. ISOLATION AND PROPERTIES OF INTACT MITOCHONDRIA FROM SPHEROPLASTS OF YEAST. J Bacteriol. 1964 Dec;88:1762–1773. doi: 10.1128/jb.88.6.1762-1773.1964. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gross S. R. The regulation of synthesis of leucine biosynthetic enzymes in Neurospora. Proc Natl Acad Sci U S A. 1965 Dec;54(6):1538–1546. doi: 10.1073/pnas.54.6.1538. [DOI] [PMC free article] [PubMed] [Google Scholar]
- NOLTMANN E. A., GUBLER C. J., KUBY S. A. Glucose 6-phosphate dehydrogenase (Zwischenferment). I. Isolation of the crystalline enzyme from yeast. J Biol Chem. 1961 May;236:1225–1230. [PubMed] [Google Scholar]
- Satyanarayana T., Umbarger H. E., Lindegren G. Biosynthesis of branched-chain amino acids in yeast: regulation of leucine biosynthesis in prototrophic and leucine auxotrophic strains. J Bacteriol. 1968 Dec;96(6):2018–2024. doi: 10.1128/jb.96.6.2018-2024.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ulm E. H., Böhme R., Kohlhaw G. Alpha-isopropylmalate synthase from yeast: purification, kinetic studies, and effect of ligands on stability. J Bacteriol. 1972 Jun;110(3):1118–1126. doi: 10.1128/jb.110.3.1118-1126.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]