Malonyl-CoA inhibition of peroxisomal carnitine octanoyltransferase

Abstract

Although the malonyl-CoA sensitivity of peroxisomal carnitine octanoyltransferase (COT) is reportedly lost on solubilization, we show that malonyl-CoA does inhibit the purified enzyme. Assay conditions such as buffer composition, pH, acyl-CoA substrate and the presence or absence of BSA can affect the observed inhibition. When assayed in the absence of BSA, COT shows simple competitive inhibition by malonyl-CoA. The Ki value for inhibition of purified COT is high (106 microM) compared with physiological concentrations (1-6 microM) and other short-chain acyl-CoA esters inhibit COT to the same degree. However, when COT is assayed in intact peroxisomes, the Ki for malonyl-CoA is almost 20-fold lower than found with the purified enzyme, whereas inhibition by other short-chain acyl-CoA esters does not change significantly. Several features of the inhibition of peroxisomal COT, including the specificity of malonyl-CoA over other short-chain acyl-CoA esters, resemble those of carnitine palmitoyltransferase (CPT)-I, suggesting that the regulation of COT and CPT-I in parallel may be necessary for the control of cellular fatty acid metabolism.

Expression of miR-33 from an SREBP2 Intron Inhibits Cholesterol Export and Fatty Acid Oxidation*

“…As mentioned before, CPT1A is the rate-limiting and most highly regulated step in ␤-oxidation. Interestingly, the dominant allosteric inhibitor malonyl-CoA regulates CROT similarly to CPT1A (28,52). In an elegant twist, evolution has maintained the regulation by miR-33 of two enzymes that are sensitive to malonyl-CoA by miR-33.…”

Role of insulin in hepatic fatty acid partitioning: emerging concepts

“…They are all immunologically distinct [16][17][18][19], and sequencing of the cDNA of several of them has so far indicated that they are separate gene products [20][21][22][23]. Interestingly, the pattern that appears to be emerging from ongoing studies is that each of these membrane systems possess two isoforms of CPT, the activity of only one of which is sensitive to malonyl-CoA inhibition [23][24][25][26][27][28][29].…”

Inhibition by etomoxir of rat liver carnitine octanoyltransferase is produced through the co-ordinate interaction with two histidine residues

“…Mutations H131A and H340A in peroxisomal COT did not have an important effect on the kinetic behaviour of the mutants with carnitine, as the differences in K m values observed for carnitine are small in either purified or peroxisomal COTs [14]. This was expected, as previous results showed that a different residue (Arg-505) of the bovine COT had a role in binding to carnitine [40].…”

“…Studies by A'Bhaird and Ramsay [14] show that malonyl-CoA binds to rat peroxisomal COT at two separate sites. At one of these sites, which is within the catalytic domain, malonyl-CoA competes with the substrate, either palmitoyl-CoA or decanoylCoA, in a similar way to the inhibition produced by several CoA esters, like glutaryl-CoA, 3-hydroxy-3-methylglutaryl-CoA and methylmalonyl-CoA.…”

“…activity by malonyl-CoA at different pHs have been reported for COT [14]. The various carnitine acyl-transferases also show differential sensitivity to inhibition by etomoxir, which is an aryl-substituted 2-oxirane carboxylic derivative [15][16][17] (Figure 1).…”