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Intracellular localization of long-chain acyl-coenzyme A hydrolase and acyl-L-carnitine hydrolase in brown adipose tissue from guinea pigs.

ABSTRACT: The activities of long-chain acyl-CoA hydrolase (palmitoyl-CoA hydrolase, EC 3.1.2.2) and long-chain acyl-L-carnitine hydrolase, EC 3.1.1.28) in brown adipose tissue from cold-exposed and control guinea pigs were studied. Mitochondria from cold-exposed animals hydrolysed 21 nmol of palmitoyl-CoA/min per mg of protein and 1.3 nmol of palmitoyl-L-carnitine/min per mg of protein, and the specific activities were respectively 2 and 5 times as high in cold-exposed as in control animals. The subcellular-localization studies showed that both the long-chain acyl-CoA hydrolase and long-chain acyl-L-carnitine hydrolase were localized in the mitochondria. A location also in the soluble fraction cannot be excluded. The long-chain acyl-CoA hydrolase activity was doubled when the mitochondria were disrupted; this indicates that the enzyme is localized in the matrix compartment.

SUBMITTER: Berge RK

PROVIDER: S-EPMC1161313 | biostudies-other | 1979 Aug

REPOSITORIES: biostudies-other

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Project description:IntroductionBrown adipose tissue (BAT) recently emerged as a potential therapeutic target in the treatment of obesity and associated disorders due to its fat-burning capacity. The current gold standard in assessing BAT activity is [18F]FDG PET-CT scan, which has severe limitations including radiation exposure, being expensive, and being labor-intensive. Therefore, indirect markers are needed of human BAT activity and volume.ObjectiveWe aimed to identify metabolites in serum that are associated with BAT volume and activity in men.MethodsWe assessed 163 metabolites in fasted serum of a cohort of twenty-two healthy lean men (age 24.1 (21.7-26.6) years, BMI 22.1 (20.5-23.4) kg/m2) who subsequently underwent a cold-induced [18F]FDG PET-CT scan to assess BAT volume and activity. In addition, we included three replication cohorts consisting of in total thirty-seven healthy lean men that were similar with respect to age and BMI compared to the discovery cohort.ResultsAfter correction for multiple testing, fasting concentrations of lysophosphatidylcholine-acyl (LysoPC-acyl) C16:1, LysoPC-acyl C16:0 and phosphatidylcholine-diacyl C32:1 showed strong positive correlations with BAT volume (β= 116 (85-148) mL, R2 = 0.81, p = 4.6 × 10-7; β = 79 (93-119) mL, R2 = 0.57, p = 5.9 × 10-4 and β= 91 (40-141) mL, R2 = 0.52, p = 1.0 × 10-3, respectively) as well as with BAT activity (β= 0.20 (0.11-0.29) g/mL, R2 = 0.59, p = 1.9 × 10-4; β = 0.15 (0.06-0.23) g/mL, R2 = 0.47, p = 2.0 × 10-3 and β= 0.13 (0.01-0.25) g/mL, R2 = 0.28, p = 0.04, respectively). When tested in three independent replication cohorts (total n = 37), the association remained significant between LysoPC-acyl C16:0 and BAT activity in a pooled analysis (β= 0.15 (0.07-0.23) g/mL, R2 = 0.08, p = 4.2 × 10-4).ConclusionsLysoPC-acyl C16:0 is associated with BAT activity in men. Since BAT is regarded as a promising tool in the battle against obesity and related disorders, the identification of such a noninvasive marker is highly relevant.

Project description:1. The nature of the acetyl-CoA hydrolase (EC 3.1.2.1) reaction in rat and sheep liver homogenates was investigated. 2. The activity determined in an incubated system was 5.10 and 3.28nmol/min per mg of protein for rat and sheep liver homogenate respectively. This activity was not affected by the addition of l-carnitine, but was decreased by the addition of d-carnitine. 3. No acetyl-CoA hydrolase activity could be detected in rat or sheep liver homogenates first treated with Sephadex G-25. This treatment decreased the carnitine concentrations of the homogenates to about one-twentieth. Subsequent addition of l-carnitine, but not d-carnitine, restored the apparent acetyl-CoA hydrolase activity. 4. Sephadex treatment did not affect acetyl-carnitine hydrolase activity of the homogenates, which was 5.8 and 8.1nmol/min per mg of protein respectively for rat and sheep liver. 5. Direct spectrophotometric assay of acetyl-CoA hydrolase, based on the reaction of CoA released with 5,5'-dithiobis-(2-nitrobenzoic acid), clearly demonstrated that after Sephadex treatment no activity could be measured. 6. Carnitine acetyltransferase (EC 2.3.1.7) activity measured in the same assay system in response to added l-carnitine was very low in normal rat liver homogenates, owing to the apparent high acetyl-CoA hydrolase activity, but was increased markedly after Sephadex treatment. The V(max.) for this enzyme in rat liver homogenates was increased from 3.4 to 14.8nmol/min per mg of protein whereas the K(m) for l-carnitine was decreased from 936 to 32mum after Sephadex treatment. 7. Acetyl-CoA hydrolase activity could be demonstrated in disrupted rat liver mitochondria but not in separated outer or inner mitochondrial membrane fractions. Activity could be demonstrated after recombination of outer and inner mitochondrial membrane fractions. The outer mitochondrial membrane fraction showed acetylcarnitine hydrolase activity and the inner mitochondrial membrane fraction showed carnitine acetyltransferase activity. 8. The results presented here demonstrate that acetyl-CoA hydrolase activity in rat and sheep liver is an artifact and the activity is due to the combined activity of carnitine acetyltransferase and acetylcarnitine hydrolase.

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Intracellular localization of long-chain acyl-coenzyme A hydrolase and acyl-L-carnitine hydrolase in brown adipose tissue from guinea pigs.

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