Microbody of n-alkane-grown yeast

Abstract

Microbodies appearing abundantly in n-alkane-grown cells of Candida tropicalis pK 233 were isolated by means of sucrose density gradient centrifugation. Electron microscopical observation showed that the microbodies isolated were intact. Localization of catalase and D-amino acid oxidase in the isolated microbodies was confirmed. Isocitrate lyase, melate synthase and NADP-linked isocitrate dehydrogenase were also located in the microbody, but malate dehydrogenase, citrate synthase, aconitase and NAD-linked isocitrate dehydrogenase were not. Neither cytochrome P-450 not NADPH-cytochrome c reductase, the components involved in the n-alkane hydroxylation system of the yeast, were detected in the microbody fraction.

Distinct subcellular localization of NAD‐linked and FAD‐linked glycerol‐3‐phosphate dehydrogenases inN‐alkane‐grownCandida tropicalis

“…Furthermore, any activities of NADH oxidation were not detected in this yeast when the TCA cycle intermediates except for oxalacetate were used individually as substrate. The activity of malate dehydrogenase was present in the cells, but not in the peroxisomes [3]. DHAP, however, was found to be effective in the oxidation of NADH by the cell-free extract of the yeast, indicating the presence of NAD-linked G3PDH in the yeast.…”

“…Thus, malate/aspartate shuttle may not operate Enzyme sources were the 3000 X g supernatant solutions of the protoplast homogenates. The enzyme activity was measured by the reduction of DHAP with NADH The experimental procedures were as in [3 ]. S~, 3000 X g supernatant of the protoplast homogenate; P2,213 000 × g pellets of S~ ; S 2, 213 000 X g supernatant of S~.…”

“…By analogy with animal mitochondria, operation of G3P/DHAP shuttle was suggested in animal peroxisomes [ 12] and malate/ aspartate shuttle (malate dehydrogenase and glutamate-oxalacetate transaminase) in plant peroxisomes [14]. Although glutamate-oxalacetate transaminase was detected in the mitochondria, peroxisomes and cytosol of alkane-grown C. tropicalis (unpublished results), malate dehydrogenase was not localized in the peroxisomes, but in the mitochondria [3]. Thus, malate/aspartate shuttle may not operate Enzyme sources were the 3000 X g supernatant solutions of the protoplast homogenates.…”

“…In the isolated peroxisomes, several enzymes were detected, e.g., catalase, isocitrate lyase, malate synthase, fatty acid/3-oxidation system, acyl-CoA synthetase and carnitine acetyltransferase [3][4][5][6]. We have suggested that peroxisomes in alkane-utilizing yeasts play essential roles in the metabolism of higher fatty acids derived from alkane substrate, i.e., fatty acid degradation and gluconeogenesis.…”

“…Measurement of NADH-cytochrome c reductase was done as that of NADPH-cytochrome e reductase [3] by using NADH in place of NADPH. NADH-glyoxylate reductase and NADH-hydroxypyruvate reductase were assayed by the methods in [7] with slight modifications.…”