Fatty acid activation

Abstract

I. Introduction A. Metabolic reactions and roles of activated fatty acids B. Enzymes involved in fatty acid activation II.Acyl-CoA synthetase reaction mechanism and assay methods A. Reactiion mechanism B. Assay methods III.Chain length specificity and species and tissue distribution of acyl-CoA synthetase activity A. Chain length specificity 1. Short-and medium-chain acyl-CoA synthetases 2. Long-chain acyl-CoA synthetase 3. Very long-chain acyl-CoA synthetase 4. Activation of unsaturated fatty acids 5. Activation of branched-chain fatty acids B. Species and tissue distribution of ACS activity IV.Organellar distribution of acyl-CoA synthetase activity A. Subcellular localization of ACS activities 1. Short-chain ACS 2. Medium-chain ACS 3. Long-chain ACS 4. Very long-chain ACS B. Very long-chain ACS activity in human peroxisomal disease C. Topog, raphic localization of ACS activity within organellar membranes V. Structural similarities between acyl-CoA synthetases and other enzymes VI.Regulation of acyl-CoA synthetase activity A. Control of enzyme activity by substrate, product, acyl-CoA binding protein, and phospholipids B. Dietary, hormonal, and substrate control of ACS expression C. Peroxisome proliferators in vivo and the regulation of long-chain ACS expression D. Short-term and in vitro effects of peroxisome proliferators on long-chain ACS activity VII.Inhibitors of acyl-CoA synthetase activity A. Carboxylic acids 1. Nonsteroidal anti-inflammatory drugs 2. Fatty acids and analogs B. Other inhibitors 1. Enoximone 2. Triacsins 3. Adenosine 5'-alkylphosphates 4. Coenzyme A analogs VIII.Activation of other carboxylic acids A. Activation of fatty acid analogs and non-fatty acid carboxylates by acyl-CoA synthetases B. Bile acid-CoA synthetase IX.Acyl-CoA regulatory functions--recent developments A. Lipid modification of proteins B. Protein kinase C activation X. Concluding remarks References exchange was dependent on acetate and CoA.Sixth, PPi and AMP were required for acetate-acetyl-CoA exchange.GTP-dependent ACS activity was also reported to be present in rat liver mitochondria, but its activity was only about 5% of the ATP-dependent activity?9~' The overall reaction mechanism, while not as well characterized as for ATP-dependent ACS, was thought to be as shown in reaction ( 4): O O II II N-C-OH + GTP + CoA-SH --* R-C-S-CoA + GDP + P~ Fatty Acid Activation 59Few reports of GTP-dependent ACS activity have appeared in the literature over the past two decades.Therefore, only the ATP-dependent enzymes will be considered in this review. B. Assay MethodsACS activity can be quantitated by several methods.For measurement of long-and very long-chain ACS activity, radioactive assays or spectrophotometric assays are commonly used.In the former, radiolabeled fatty acids are incubated with ATP/Mg ÷ + and CoA in the presence of ACS.Labeled acyl-CoA product can be separated from the substrate either by partition using the method of Dole 62' 63 or Folch, ~4' 65 by reverse-phase HPLC separation, 66 or by capillary electrophoresis.67 Partition methods must be used with care when assaying activation of short-and medium-chain fatty acids because these substrates are more soluble in aqueous medium than are long-or very long-chain fatty acids.ACS activity can also be assayed by monitoring directly the formation of the thioester bond at 232 nm.68' 69 In addition, several coupled assays for measurement of long-chain ACS activity have been described.Tanaka and coworkers 7° used an assay in which AMP formation was coupled to lactate dehydrogenase via adenylate kinase and phosphoenolpyruvate; oxidation of NADH was measured at 340 nm.More recently, assays have been described in which the acyl-CoA product is oxidized via acyl-CoA oxidase, yielding H202 which is then coupled via peroxidase either to oxidation of a dye detectable at 550 nm 71, or to formation of a fluorescent compound.72Other methods for assessing ACS activity are less commonly used.Acyl-CoA formed in the reaction can be trapped either as the hydroxamate 43 or as an acyl-carnitine.73 Bar-Tana and coworkers described assays measuring either CoA sulfhydryl disappearance TM or AMP formation75; measurement of PP~ release has also been utilized.43 III. CHAIN LENGTH SPECIFICITY AND SPECIES AND TISSUE DISTRIBUTION OF ACYL-COASYNTHETASEACTIVITY A. Chain Length SpecificityACSs are generally classified by their specificities for fatty acids of varying chain length.The Commission on Enzymes of the International Union of Biochemistry has classified these enzymes as acetyl-CoA synthetase (EC 6.2.1.1)medium-chain ACS (EC 6.2.1.2) and long-chain ACS (EC 6.2.1.3)However, there is generally considerable overlap in chain length specificity between different enzymes.This overlap has led to difficulty in interpreting results obtained in crude systems.However, by studying both enzymes purified by conventional methods and enzymes expressed in vitro, researchers have begun to understand their sp