Subcellular Localization of the Leucine Biosynthetic Enzymes in Yeast

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Issue Date
1973-10Author
Ryan, Eric D.
Tracy, James W.
Kohlhaw, Gunter B.
Publisher
American Society for Microbiology
Type
Article
Article Version
Scholarly/refereed, publisher version
Published Version
http://jb.asm.org/content/116/1/222.abstract?sid=29df9b7c-1907-4a98-9a13-712500e03f97Metadata
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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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Citation
Ryan, E. D., Tracy, J. W., & Kohlhaw, G. B. (1973). Subcellular Localization of the Leucine Biosynthetic Enzymes in Yeast. Journal of Bacteriology, 116(1), 222–225.
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