Biochemical Competition Makes Fatty-Acid β-Oxidation Vulnerable to Substrate Overload

Abstract

Fatty-acid metabolism plays a key role in acquired and inborn metabolic diseases. To obtain insight into the network dynamics of fatty-acid β-oxidation, we constructed a detailed computational model of the pathway and subjected it to a fat overload condition. The model contains reversible and saturable enzyme-kinetic equations and experimentally determined parameters for rat-liver enzymes. It was validated by adding palmitoyl CoA or palmitoyl carnitine to isolated rat-liver mitochondria: without refitting of measured parameters, the model correctly predicted the β-oxidation flux as well as the time profiles of most acyl-carnitine concentrations. Subsequently, we simulated the condition of obesity by increasing the palmitoyl-CoA concentration. At a high concentration of palmitoyl CoA the β-oxidation became overloaded: the flux dropped and metabolites accumulated. This behavior originated from the competition between acyl CoAs of different chain lengths for a set of acyl-CoA dehydrogenases with overlapping substrate specificity. This effectively induced competitive feedforward inhibition and thereby led to accumulation of CoA-ester intermediates and depletion of free CoA (CoASH). The mitochondrial [NAD+]/[NADH] ratio modulated the sensitivity to substrate overload, revealing a tight interplay between regulation of β-oxidation and mitochondrial respiration.

Measurement of β-oxidation capacity of biological samples by respirometry: a review of principles and substrates

“…Carnitine scavenges CoA esters and replenishes CoA, as discussed earlier (see Fig. 5) (20,69). Bressler and Katz (10) and others (18,19) have demonstrated using whole liver homogenates and isolated liver mitochondria from guinea pigs that in the fasted state carnitine stimulates fatty acid oxidation and acetoacetate formation by stimulating 1) transport of long-chain fatty acyl groups into the mitochondria for oxidative conversion to acetoacetyl-CoA and 2) acetoacetylcarnitine formation and transport out of the mitochondria for acetoacetate production.…”

“…However, their effective oxidation requires that malate or carnitine be added as cosubstrates (36). Van Eunen et al (69) recently showed that the cyclic ␤-oxidation pathway results in two types of competition: 1) substrates of different chain length compete for common enzymes, and 2) enzymes with overlapping chain length specificity compete for common substrates. These competitions lead to a feedforward inhibition in the network and accumulation of shorter-chain acyl-CoAs that contribute to the depletion of matrix CoA as well as a buildup of acetyl-CoA and NADH that inhibits the thiolase reaction of ␤-oxidation and decreases oxygen flux (37,57).…”

“…Oxidation of malate to oxaloacetate produces NADH beyond ␤-oxidation (23), and condensation of oxaloacetate with acetyl-CoA produces citrate, which appears not to be metabolized further because it accumulates in the oxygraph media and accounts for most of the carbon atoms consumed in palmitoylcarnitine oxidation (69). Indeed, it is known that the tricarboxylate carrier found on the inner mitochondrial membrane is very effective in transporting citrate out of the mitochondrial matrix in exchange for malate (see Fig.…”

“…Therefore, the measured oxygen flux is due to electrons from one NADH from malate dehydrogenase, one NADH from enoyl-CoA dehydrogenase, and one FADH 2 from acyl-CoA dehydrogenase. Since each O 2 oxidizes two NADH or FADH 2 , the oxygen flux from ␤-oxidation per se corresponds to two-thirds of the measured flux (69).…”

Drug target identification through systems biology

“…www.drugdiscoverytoday.com e19 models exist for glycolysis of T. brucei ( [26], most recent version in [27]), yeast [28], erythrocytes [29] and liver [30], but also other relevant pathways have been modelled kinetically, such as glycogenolysis in skeletal muscle [31], b-oxidation of fatty acids [32] and gluthatione metabolism [33] in liver cells. Metabolic control analysis (reviewed in [34]), a method to quantify which enzyme, when partially reduced in activity, has the strongest effect on a pathway flux, revealed that the glucose uptake into T. brucei exerted the highest control over the vital glycolytic flux of this parasite [35] ( Fig.…”