Rat Long Chain Acyl-CoA Synthetase 5, but Not 1, 2, 3, or 4, Complements Escherichia coli fadD

Abstract

Long chain fatty acids are converted to acyl-CoAs by acyl-CoA synthetase (fatty acid CoA ligase: AMP forming, E.C. 6.2.1.3; ACS). Escherichia coli has a single ACS, FadD, that is essential for growth when fatty acids are the sole carbon and energy source. Rodents have five ACS isoforms that differ in substrate specificity, tissue expression, and subcellular localization and are believed to channel fatty acids toward distinct metabolic pathways. We expressed rat ACS isoforms 1-5 in an E. coli strain that lacked FadD. All rat ACS isoforms were expressed in E. coli fadD or fadDfadR and had ACS specific activities that were 1.6 -20-fold higher than the wild type control strain expressing FadD. In the fadD background, the rat ACS isoforms 1, 2, 3, 4 and 5 oxidized [ 14 C]oleate at 5 to 25% of the wild type levels, but only ACS5 restored growth on oleate as the sole carbon source. To ensure that enzymes of ␤-oxidation were not limiting, assays of ACS activity, ␤-oxidation, fatty acid transport, and phospholipid synthesis were also examined in a fadD fadR strain, thereby eliminating FadR repression of the transporter FadL and the enzymes of ␤-oxidation. In this strain, fatty acid transport levels were low but detectable for ACS1, 2, 3, and 4 and were nearly 50% of wild type levels for ACS5. Despite increases in ␤-oxidation, only ACS5 transformants were able to grow on oleate. These studies show that although ACS isoforms 1-4 variably supported moderate transport activity, ␤-oxidation, and phospholipid synthesis and although their in vitro specific activities were greater than that of chromosomally encoded FadD, they were unable to substitute functionally for FadD regarding growth. Thus, membrane composition and proteinprotein interactions may be critical in reconstituting bacterial ACS function.

Promotion of glioma cell survival by acyl-CoA synthetase 5 under extracellular acidosis conditions

“…We have shown earlier that inhibiting multiple ACS activities strongly induces apoptosis, whereas this cell death is almost completely suppressed by a single gene transfer of ACSL5 . In addition, among mammalian ACS, only ACSL5 restores the growth of an Escherichia coli strain that lacks FadD, the only known ACS enzyme in the E. coli (Caviglia et al, 2004). These observations suggest that among ACS members, ACSL5 could have a predominant function in cell survival.…”

Mammalian Long-Chain Acyl-CoA Synthetases

“…Therefore, the activity of these enzymes will be affected by both lipid-lipid, lipidprotein and protein-protein interactions at the membrane water interface (19,33). A number of studies of the activity of the different forms of this family of enzymes have been reported, but we argue that the majority of such data should be evaluated very carefully.…”

“…The activity of these enzymes is strongly dependent on their lipid environment. The presence of lipids enhance their activity and the lipid composition of the membranes in which the enzymes are measured has been reported to affect their activity (19,33). Rescue of acyl-CoA synthetase mutants by the mammalian forms were performed in both E. coli and S. cerevisiae strains.…”

Long-Chain Acyl-Coa Synthetases And Fatty Acid Channeling

“…Unique roles for ACS isoforms were first identified in studies of Saccharomyces cerevisiae mutants that lacked the ACS isoforms Faa1 and Faa4, and were unable to use exogenously provided fatty acids [31]. Similarly, complementation studies of ACS-deficient E. coli showed that each of the 5 rat ACSL isoforms differs in its ability to channel FA into specific pathways like phospholipid synthesis and β-oxidation [32]. The ACSL and FATP isoforms also differ in their ability to complement FA uptake and activation in mutated yeast strains that lack ACS activity [28,33].…”