Abstract
Fatty acid synthetase purified (20 times) from lactating bovine mammary tissue had an approximate mot wt of 485,000. The enzyme had a high content of acidic and hydrophobic amino acid residues; 62±4 sulhydryl groups and one 4′ phosphopantetheine residue/mole of enzyme. The enzyme was relatively stable when stored (3 mg/ml) in potassium phosphate buffer (250 mM), containing dithiothreitol (5 mM) at −5 C or at −30C or as a lyophilized powder at −30 C. Preincubation at 37 C in presence of dithioltreitol (5 mM) was necessary for obtaining maximum activity at the optimum pH of 6.8. Maximum specific activity of the isolated enzyme was 55 nmoles acetyl-coenzyme A min−1mg−1 incorporated A min−1mg−1 incorporated into fatty acids. Butyryl-coenzyme A or acetyl-coenzyme A (30μM), malonyl-coenzyme A (65 μM), and nicotinamide adenine dinucleotide phosphate, reduced form (300 μM) were required for optimum fatty acid synthesis. Malonyl-coenzyme A decarboxylase activity (5 nmoles min−1mg−1) associated with the enzyme permitted fatty acid synthesis in the presence of nicotinamide adenine dinucleotide phosphate, reduced form and malonyl-coenzyme A. The enzyme utilized acetyl-coenzyme A, butyryl-coenzyme A, and hexanoyl-coenzyme A as primers, with butyryl-coenzyme A giving the maximum rate of fatty acid synthesis. Apparent Km values of 22,6.7, 3, 22, and 20 μM were obtained for malonyl-coenzyme A, acetyl-coenzyme A, butyryl-coenzyme A, hexanoyl-coenzyme A, and nicotinamide adenine dinucleotide phosphate, reduced form, The fatty acid synthetase was inhibited by N-ethylmaleimide, iodoacetamide, and progressively inhibited by increasing concentrations of long chain acyl-coenzyme A, i.e. palmityl-coenzyme A and myristyl-coenzyme A. This inhibition was relieved by bovine serum albumin or β-lactoglobulin (3 mg/ml). Palmitic acid was the major product of bovine mammary fatty acid synthetase. However, small amounts of fatty acids, 4∶0–14∶0 inclusive, also were synthesized. The pattern of fatty acids was altered by varying malonyl-coenzyme A to acetyl-coenzyme A ratios and by increasing the enzyme levels in the assays. At high concentrations of enzyme (0.5 mg/ml), greater amounts of short and medium chain fatty acids were generated.
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Kinsella, J.E., Bruns, D. & Infante, J.P. Fatty acid synthetase of bovine mammary: Properties and products. Lipids 10, 227–237 (1975). https://doi.org/10.1007/BF02532485
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DOI: https://doi.org/10.1007/BF02532485