Isolation of a novel type-I fatty-acid synthetase from Euglena gracilis. Specific derepression in streptomycin-bleached cells.
A novel, high-molecular-mass fatty-acid synthetase (FAS) complex has been isolated from streptomycin-bleached Euglena gracilis cells. The enzyme was purified 250-fold from the crude cell homogenate and subsequently migrated upon SDS/PAGE as a single band of molecular mass 270 kDa. This apparent subunit size of the purified protein contrasted with a smaller size of only 200 kDa which was exhibited by the same protein upon immunoblotting of the crude cell extract. The purified Euglena FAS complex cosediments in a sucrose density gradient with yeast FAS and, from this, both enzymes were concluded to have the same overall molecular mass of 2.3 MDa. The enzyme described in this paper appears to be a typical type-I FAS multienzyme which clearly differs from the E. gracilis FAS so far described. Instead, it appears to be organized structurally similar to the type-I FAS multienzymes of lower fungi. In vitro, the purified Euglena FAS complex synthesizes mainly palmitic acid, or its CoA ester, from acetyl CoA and malonyl CoA as substrates. The Km values for acetyl CoA and malonyl CoA are 20 microM and 31 microM, respectively. Similar to the FAS enzymes of other lower eucaryotes, the Euglena type-I FAS is a flavoprotein. In contrast to yeast FAS, however, the flavin cofactor appears to be covalently attached to the enzyme protein. By immunological techniques, the enzyme was shown to be absent in green as well as in etiolated E. gracilis cells, while being rapidly induced upon streptomycin bleaching of heterotrophically growing green cells. The data suggest an inverse correlation between organellar development and derepression of this FAS complex.
Siebenlist
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U
Lehrstuhl für Biochemie, Universität Erlangen-Nürnberg, Federal Republic of Germany.
Wohlgemuth
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Finger
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K
Schweizer
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E
D000818:Animals; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D005056:Euglena gracilis / Q000187:drug effects / Q000201:enzymology*; D064429:Fatty Acid Synthases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D005453:Fluorescence; D013307:Streptomycin / Q000494:pharmacology
D064429:Fatty Acid Synthases; D013307:Streptomycin