Distribution of reaction intermediates on chicken liver fatty acid synthase.
The distributions of covalent intermediates in the reaction cycle catalyzed by chicken liver fatty acid synthase were studied. In isotope-trapping experiments, 30% of [1-14C] acetyl-enzyme and 6.7% of [2-14C]malonyl-enzyme are converted to long-chain fatty acids, indicating the initiation reaction is partially random. The 3-hydroxybutyryl intermediate is located on the tryptic peptide which contains the 4'-phosphopantetheine (greater than 90%), while the C4-C18 saturated intermediates are distributed both on this peptide and on the peptide that contains the active cysteine. The ratio of intermediates on the two peptides is about unity for chain lengths less than C14, but the amount on the active cysteine progressively decreases for chain lengths of C14, C16, and C18 with trypsinized enzyme. The distributions of carbon chain lengths for the saturated or 3-keto intermediates when acetoacetyl-labeled trypsinized enzyme is incubated with limiting malonyl coenzyme A or NADPH, respectively, show large fractions both of unreacted enzyme and of C16 or longer intermediates. A detailed analysis suggests that the initial condensation and reduction steps are slower than the analogous reactions with longer chain length intermediates. The 3-keto intermediate comprises over 70% of each chain length intermediate detected when NADPH is the limiting substrate, indicating the reduction of the 3-keto intermediates is at least 2 times slower than the reduction of the unsaturated intermediates.
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- Trypsin
- NADP
- Models, Chemical
- Malonyl Coenzyme A
- Liver
- Fatty Acid Synthases
- Chromatography, High Pressure Liquid
- Chickens
- Biochemistry & Molecular Biology
- Animals
Citation
Published In
DOI
ISSN
Publication Date
Volume
Issue
Start / End Page
Location
Related Subject Headings
- Trypsin
- NADP
- Models, Chemical
- Malonyl Coenzyme A
- Liver
- Fatty Acid Synthases
- Chromatography, High Pressure Liquid
- Chickens
- Biochemistry & Molecular Biology
- Animals