Biochemical

and Molecular Roles of Nutrients

Rat Hepatic Coenzyme A Is Redistributed in Response to Mitochondrial Acyl-Coenzyme A Accumulation1»2 Departments of Medicine and Pharmacology, Diuision of Clinical Pharmacology, Case Western Reserve University, Cleveland, OH 44106 substrate recognition. Because the cell is dependent on the continual availability of CoASH, low cellular total CoA content (CoASH plus all acyl-CoAs) or sequestration of CoASH as poorly metabolized acylCoAs will impair cellular metabolism (Chalmers et al. 1983). Coenzyme A is synthesized in all cells from pantothenate, cysteine and ATP (Robishaw and Neely 1985). The regulated, rate-limiting step in CoA bi osynthesis is phosphorylation of pantothenate by pantothenate kinase in the cytosol (Robishaw et al. 1982, Vallari et al. 1987). Terminal steps in the biosynthesis of CoA can occur in both the cytosol and mitochondria. In contrast, CoA degradation seems to be exclusively cytosolic (Bremer et al. 1972, Skrede 1973). Coenzyme A-requiring reactions take place in both the cytosol and mitochondria, and movement of CoA across the inner mitochondrial membrane occurs via a specific transport system (Tahiliani and Neely 1978). Coenzyme A sequestration by acyl-CoAs results in enhanced CoA biosynthesis (Brass et al. 1990a, Roitman and Smith 1987). This increased bi osynthesis, and resultant increased total cellular CoA, may be an important compensatory mechanism for normalizing metabolism under conditions of acylCoA accumulation (Brass 1992, Brass et al. 1990a, Russell and Taegtmeyer 1992, Veitch et al. 1987). For example, hepatocytes from clofibrate-treated rats

ABSTRACT Coenzyme A without an acyl-thioester (CoASH) is required for numerous cellular reactions, and sequestration of CoASH as acyl-CoAs may impair metabolic function. Increased total CoA protects the cell from acyl-CoA accumulation, and enhanced CoA bi osynthesis may represent a compensatory response in metabolic disease. To test the hypothesis that cellular CoA is redistributed from the cytosol to the mitochondria in response to mitochondrial acyl-CoA accretion, the subcellular distribution of hepatic CoA was determined by differential centrifugation and meas urement of the mitochondrial marker enzyme citrate synthase. Liver from control, clofibrate-treated and hydroxycobalamin[c-lactam] (HCCL)-treated rats were used. Clofibrate increased total hepatic CoA concen tration 2.2-fold, whereas HCCL (which causes inhibition of L-methylmalonyl-CoA mutase and consequent propionyl- and methylmalonyl-CoA accumulation) in creased it threefold. However, clofibrate did not affect the percentage of total CoA in the mitochondria (control: 44 ±3%, clofibrate: 49 ±5%), and HCCL-treatment induced a marked redistribution of CoA into the mitochondria (HCCL: 78 - 8%). Redistribution of total CoA was also induced acutely by incubation of hepatocytes from control rats with 10 mmol/L propionate. Thus, redistribution of the cellular CoA pool can help maintain CoASH availability as mitochondrial acyl-CoA accumulation occurs and may be an important compen satory response to metabolic injury. J. Nutr. 122: 2094-2100, 1992. INDEXING KEY WORDS:

•rats •coenzyme A •cobalamin •propionate •clofibrate

1Presented at 1992 FASEB meeting, Anaheim, CA (Brass, E. P. &. Ruff, L. y. (1992| Redistribution of hepatic coenzyme A in response to mitochondrial acyl-CoA accumulation. FASEB J. 6: A1519 (abs.)]. ^his work was supported by NIH DK36069. Eric Paul Brass is a Burroughs Wellcome Scholar in Clinical Pharmacology. Abbreviations used: CoASH, coenzyme A without an acylthioester; HCCL, hydroxycobalaminjc-lactam]; MSM, buffer con taining 220 mmol/L mannitol, 70 mmol/L sucrose and 5 mmol/L 3-[N-morpholino]propanesulfonic acid.

Coenzyme A without an acyl-thioester (CoASH)3 is an important cofactor for a broad spectrum of biosynthetic and oxidative reactions in mammalian cells. The energy-dependent formation of an acyl-CoA thioester acidifies the ct-ßhydrogens of the organic acid, facilitating subsequent transformations (Putter 1957). The CoA moiety also contributes to enzyme 0022-3166/92

$3.00 ©1992 American

Institute

of Nutrition.

Received 10 April 1992. Accepted 26 June 1992.

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ERIC P. BRASS AHD LAURA J. RUFF

REDISTRIBUTION

MATERIALS

AND METHODS

Animals and hepatocyte isolation. All procedures involving animals were reviewed and approved by the Case Western Reserve University Animal Welfare Committee. Male Sprague-Dawley rats (Zivic-Miller, Allison Park, PA) were allowed free access to nonpurified diet (Formulab Chow 5008, Purina, Richmond, IN) and water and studied in the fed state. Rats weighed 380 ±80 g (n = 21) when experiments were initiated. Clofibrate-fed rats were fed the nonpurified diet supplemented with 0.5% (wt/wt) clofibrate for 9-11 d. Both control and clofibrate-treated rats were maintained in individual wire-bottom cages. Food consumption and weight gain is not altered by this clofibrate-treatment protocol (Brass and Ruff 1991). Methylmalonic aciduria secondary to cobalamin deficiency-associated decreased L-methylmalonylCoA muÃ-ase activity was induced by 2-wk sub cutaneous treatment with hydroxycobalaminjclactam] (HCCL) (2 jig/h) delivered by osmotic minipump (Alza, Palo Alto, CA, Model 2002) as previ ously described (Brass et al. 1990b, Krahenbuhl et al. 1990). Hepatocytes were isolated from fed, control rats by the collagenase perfusion technique of Berry and

2095

Friend ¡1969)as previously detailed (Brass and Ruff 1991). Hepatocyte preparations used in the current studies averaged 97 ±1% viable based on Trypan Blue exclusion (12= 7), contained 6.0 ±0.8 mg protein/IO6 cells (n = 7) and had a wet weight of 22.4 ±6.3 mg/106 cells (n = 4). Hepatocytes (8.14 ±0.68 x IO9 cells/L, n = 7) were incubated in 126 mmol/L NaCl, 3.76 mmol/ L KC1, 1.90 mmol/L MgSO4, 0.95 mmol/L KH2PO4, 20 mmol/L NaHCO3, 0.8 mmol/L glucose, 1.1 mmol/ L CaCl2, 10 g/L defatted bovine serum albumin at 37°C, pH 7.4 under 95% O2-5% CO2. Following a 10-min preincubation, propionate or diluent was added as indicated in individual experiments. Subcellular distribution. To permit calculation of the subcellular distribution of hepatic CoA, highly purified mitochondria were prepared by differential centrifugation by a modification of the method of Hoppel et al. (1979). Rats were killed by decapitation, and livers were rapidly excised, rinsed in cold 220 mmol/L mannitol, 70 mmol/L sucrose, 5 mmol/L 3-[AT-morpholino]propanesulfonic acid, pH 7.4 (MSM) and washed with MSM (all subsequent steps at 4°C). A portion of the liver (6-8 g) was then minced and homogenized in MSM containing 8 mmol/L EDTA using a Potter-Elvehjem glass/teflon homogenizer. An aliquot of the homogenate was saved for analysis, and the remainder centrifuged at 700 x g for 10 min. The resulting supernatant was sampled ("E" fraction), and centrifuged at 7600 x g for 10 min to pellet mitochondria. This pellet, containing the heavymitochondrial and light-mitochondrial fractions (Hoppel et al. 1979, DeDuve et al. 1955), was vigorously shaken and wiped with gauze to remove the light-mitochondrial fraction (mitochondria, peroxisomes and lysosomes). The mitochondrial pellet was resuspended in MSM and washed twice using the above procedure to yield the final mitochondrial fraction ("M"). All 7600 x g supernatants were pooled to yield the "S" fraction. The 700 x g pellet was resuspended in MSM as the "N" fraction. The removal of the light-mitochondrial fraction decreased recoveries (see Results and Discussion), but assured high purity mitochondrial fractions. Coenzyme A in the mitochondrial fraction was used to determine mitochondrial vs. cytosolic CoA distribution based on the mitochondrial marker enzyme citrate synthase: fraction of total CoA in mitochondria = (total CoA in M -s-fraction of total citrate synthase in M) -i- (total CoA in H), where fraction of total citrate synthase in M = citrate synthase in M + citrate synthase in H. Coenzyme A distribution in hepatocytes was esti mated following digitonin fractionation (Zuurendonk and Tager 1974). An aliquot of the hepatocyte incu bation (288 uL) was added to 962 uL of digitonin medium (250 mmol/L sucrose, 20 mmol/L MOPS, 3 mrnol/L EDTA, 6.5 g/L digitonin, pH 7.0). The hepatocyte-digitonin mixture was allowed to sit on ice for 20 s, and then centrifuged at 10,000 x g for 20

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contain two- to threefold more total CoA as compared with cells from control rats, and are comparatively resistant to inhibition of pyruvate oxidation by propionyl-CoA and methylmalonyl-CoA accretion (Brass 1992). Clofibrate treatment is also partially protective against hypoglycin toxicity (Veitch et al. 1987). Critical to this protective effect of CoA availa bility is the maintenance of CoASH concentrations in the cellular compartment in which the acyl-CoA ac cumulation occurs (i.e., intramitochondrial in the case of methylmalonyl-CoA build-up from exogenous propionate). Thus, although increased total cellular CoA content is important, proper cellular distribution of the CoA is also implicitly necessary. This redistri bution of the cellular CoA pool into the mitochondria might be facilitated by depletion of matrix CoASH as acyl-CoAs are formed, maintaining a trans-mitochondrial membrane CoASH gradient. Cellular redistri bution of total CoA with altered metabolic state has been demonstrated in heart (Lopaschuk and Neely 1987). The current studies were undertaken to challenge the hypothesis that mitochondrial acyl-CoA accumu lation will result in a redistribution of total cellular CoA from the cytosol into the mitochondria. This hypothesis was evaluated in vivo in a model of chronic mitochondrial propionyl-CoA and methylmalonyl-CoA accumulation (cobalaminanalog-induced hepatic cobalamin deficiency) and acutely, in vitro using hepatocytes incubated with propionate.

OF HEPATIC CoA

2096

BRASS AND RUFF

CoAs

= LDHS LDH

x CoAcytosol

CSS

+ cs~

X ^°^nuto

CoAp = LDHp CoAcytosol

+ C&; X CoAmito

where CoA$, LDH$, and CSs are the total CoA con centrations, lactate dehydrogenase and citrate syn thase activities in the digitonin-supernatant, respec tively. Similarly, CoAp, LDHp and CSp refer to the three variables in the digitonin-pellet, and LDH-p and CSj are the total (supernatant plus pellet) activities. CoAcytosoi and CoAm¡toare the actual cytosolic and mitochondrial total CoA concentrations and are cal culated based on the actual measurement of all other variables in the two equations. Assays. Coenzyme A was measured using the fluorometric enzyme recycling assay developed by Allred and Guy (1969) as previously described (Brass and Beyerinck 1988). Total acid-soluble CoA (CoASH plus acyl-CoAs of acyl-group chain length 0.05) following incubation with propionate. This propionate-induced redistri bution was observed despite only a 1-h incubation period and a higher initial mitochondrial CoA content in the hepatocytes as compared with in vivo liver. Similar results were obtained using succinate de hydrogenase as a mitochondrial marker enzyme (data not shown). Thus, mitochondrial acyl-CoA accumu lation is associated with an acute redistribution of the cellular CoA pool. Cellular CoA homeostasis includes distribution be tween CoASH and acyl-CoAs, CoA degradation and biosynthesis, and the subcellular distribution of the CoA pool. Clofibrate treatment induces pantothenate kinase activity, the rate-limiting step in CoA bi osynthesis, without perturbation of the CoASH-acylCoA distribution. As a result, as total CoA per gram of liver is increased, the subcellular distribution mimics that in the control state (Table 2). In contrast, HCCL treatment induces marked acyl-CoA accumu lation (Brass et al. 1990a). Further, this accumulation is predominantly intramitochondrial due to the mitochondrial localization of L-methylmalonyl-CoA mutase (Fenton et al. 1984). Acyl-coenzyme A ac cumulation can acutely increase CoA synthesis from pantothenic acid (Brass et al. 1990a, Roitman and Smith 1987). In the HCCL-treated rat, as the total cellular CoA content increases, the CoA is preferen tially localized to the mitochondria (Table 2) as matrix CoASH is sequestered as propionyl- and

REDISTRIBUTION

OF HEPATIC CoA

2099

TABLE 3

TABLE 4

Digitonin fractionation of rat bepatocytes incubated in the ab sence or presence of propionate

Propionate-induced redistribution of the coenzyme A pool in rat bepatocytes1

Control I" - 7) cells)

In S, % In P, % Láclate dehydrogenase Total, \imol/(min-106

cells)

In S, % In P, % Coenzyme A & acyl-CoA Total, nmol/106 cells In S, % In P, %

158 10 90

±17 ± 4 ± 4

157 92

±17 ± 1 ± 1

8.94 ± 1.11 87 ± 5 13 ± 5

9.04 ± 1.26 87 ± 4 13 ± 4

6.17 ± 0.91 21 ± 2 79 ± 2

6.37 ± 1.09 16 ± 2 84 ± 2

'Hepatocytes were isolated from control rats and incubated in the absence (control) or presence of 10 mmol/L propionate for 60 min. Following incubation, subcellular fractionation was performed by digitonin treatment yielding a pellet (P) and supernatant (S) which were assayed for citrate synthase, lactate dehydrogenase and total CoA. Total values refer to the sum of S + P. Values are means ±SEM.

methylmalonyl-CoA. As pantothenate kinase is a cytosolic enzyme and is inhibited by CoASH (Abiko et al. 1972, Fisher and Neely 1985), acute redistri bution of CoA from the cytosol to mitochondria may contribute to HCCL-induced enhanced CoA bi osynthesis. This concept is supported by the similar cytosolic total CoA concentrations in the control (160 nmol/g) and HCCL-treated (190 nmol/g) rat liver, despite the threefold difference in hepatic total CoA concentrations. The utility of citrate synthase to characterize mitochondrial distribution during fractionation in the current studies is potentially limited secondary to the differential molecular weight of this protein and CoA. However, if substantial loss of CoA from the mitochondria occurred during isolation, the marked HCCL-induced mitochondrial accumulation would not have been observed. Additional factors supporting the concept that minimal mitochondrial CoA loss has occurred are the high oxidative capacity and respiratory control ratios for mitochondria isolated by this method (Hoppel et al. 1979), and the main tenance of a mitochondrial-cytosolic total CoA con centration gradient based on the observed distribution (hepatic mitochondrial volume is -20% of total hepatic volume). Maintaining CoASH availability in both the cyto solic and mitochondrial compartments is essential for normal cellular intermediary metabolism. Under a variety of pathophysiologic conditions characterized by unusual acyl-CoA accumulation, increased total

Control(73 -6)Percentage

-

in:Mitochondria, of hepatocyte %Cytosol, %Total

CoA

in:Mitochondria, CoA content cellsCytosol, nmol/10^

nmol/106 cells77 'Data from digitonin-fractionation

(n

7)Propionate

±323 ±44.70

2*16 ± 2*5.39±

0.751.32 ± ±0.2184 of hepatocytes

0.981.01 ± ±0.16 presented in

Table 3 was used to calculate the mitochondrial and cytosolic total coenzyme A contents as described in Materials and Methods. Values are means ±SEM. *P < 0.05 vs. control.

CoA content may mitigate toxicity (Brass 1992, Brass et al. 1990a, Russell and Taegtmeyer 1992, Veitch and van Hoof 1990, Veitch et al. 1987). For example, the hepatocytes from clofibrate-treated rats are relatively resistant to propionate inhibition of pyruvate toxicity (Brass 1992). Propionate toxicity is not well predicted based solely on the propionyl-CoA content, but is dependent on CoASH depletion as well (Brass 1992). Similarly, clofibrate treatment is protective against hypoglycin (Veitch et al. 1987) and valproate (Veitch and van Hoof 1990) toxicities, other organic acids de pendent on acyl-CoA generation for affect (Ito et al. 1990, Schulz 1987). However, a protective effect of increased hepatic total CoA is dependent on increased CoA in the mitochondrial compartment. The current studies confirm the increased mitochondrial availa bility of CoA in both the clofibrate- and HCCLtreated rats. Increased total cellular and mitochondrial CoA concentrations may be an important compensatory mechanism in metabolic disease. Increased hepatic CoA in the cobalamin-deficient methylmalonic aciduria rat model may contribute to relatively normal fuel homeostasis in vivo (Brass et al. 1990a). Similar mechanisms may be important in the human organic acidurias.

ACKNOWLEDGMENTS The authors thank Charles Hoppel for his con structive comments on the manuscript. We also thank F. S. Leeds for assisting in the synthesis of hydroxycobalamin[c-lactam] used in the studies.

LITERATURECITED Abiko, Y., Ashida, S. I. & Shimizu, M. (1972) Purification and properties of D-pantothenate kinase from rat liver. Biochim.

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Citrate synthase Total, nmol/imin-lO6

Propionate (n -6|

2100

BRASS AND RUFF coenzyme A degradation by fatty acids. Am. J. Physiol. 253: H41-H46. Lowry, O. H., Rosebrough, N. J., Farr, L. &. Randall, R. J. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193: 265-275. Lund, H., Stakkestad, J. A. & Skrede, S. (1986) Effects of thyroid state and fasting on the concentrations of CoA and malonylCoA in rat liver. Biochim. Biophys. Acta 876: 685-687. Michal, G. & Bergmeyer, H. U. (1974) Determination of CoA-SH with phosphotransacetylase. In: Methods of Enzymatic Analysis (Bergmeyer, H. U., éd.),pp. 1972-1975. Academic Press, New York, NY. Putter, J. (1957) Überden Wirkungsmechanismus von coenzym A. Z. Physiol. Chem. 308: 81-90. Robishaw, J. D., Berkich, D. & Neely, f. R. (1982) Rate limiting step and control of coenzyme A synthesis in cardiac muscle. J. Biol. Chem. 257: 10967-10972. Robishaw, J. D. & Neely, I. R. (1985) Coenzyme A metabolism. Am. J. Physiol. 248: E1-E9. Roitman, K. J. &. Smith, C. M. (1987) Stimulation of CoA bi osynthesis by p-aminobenzoate and benzoate in cultured rat hepatocytes. Fed. Proc. 46: 2263. Russell, R. R. III. & Taegtmeyer, H. (1992) Coenzyme A seques tration in rat hearts oxidizing ketone bodies. J. Clin. Invest. 89: 968-973. Schulz, H. (1987) Inhibitors of fatty acid oxidation. Life Sci. 40: 1443-1449. Skrede, S. (1973) The degradation of CoA: subcellular localization and kinetic properties of CoA- and dephospho-CoA pyrophosphatase. Eur. f. Biochem. 38: 401-407. Skrede, S. & Halvorsen, O. (1979) Increased biosynthesis of CoA in the liver of rats treated with clofibrate. Eur. J. Biochem. 98: 223-229. Smith, C. M., Cano, M. L. & Potyraj, J. (1978) The relationship between metabolic state and total CoA content of rat liver and heart. J. Nutr. 108: 854-862. Srere, P. A. (1969) Citrate synthase. Methods Enzymol. 13: 3-11. Stabler, S. P., Brass, E. P., Marceli, P. D. & Allen, R. H. (1991) Inhibition of cobalamin-dependent enzymes by cobalamin ana logues in rats. J. Clin. Invest. 87: 1422-1430. Tahiliani, A. G. & Neely, J. R. (1987) A transport system for coenzyme A in isolated rat heart. J. Biol. Chem. 262: 11607-11610. Tandler, B., Krahenbuhl, S. & Brass, E. P. (1991) Unusual mitochondria in the hepatocytes of rats treated with a vitamin Bj2 analogue. Anat. Ree. 231: 1-6. Vallari, D. S., Jackowski, S. & Rock, C. O. (1987) Regulation of pantothenate kinase by coenzyme A and its thioesters. J. Biol. Chem. 262: 2468-2471. Vassault, A. (1983) Lactate dehydrogenase. In: Methods of En zymatic Analysis (Bergmeyer, H. U., ed.), vol. ffl, pp. 118-125. Verlag Chemie GmbH, Weinheim, Germany. Veitch, K. & van Hoof, F. (1990) In vitro effects of eight-carbon fatty acids on oxidations in rat liver mitochondria. Biochem. Phar macol. 40: 2153-2159. Veitch, R. K., Sherratt, H.S.A. & Bartlett, K. (1987) Organic aciduria in rats made resistant to hypoglycin toxicity by pretreatment with clofibrate. Biochem. J. 246: 775-778. Zuurendonk, P. F. & Tager, J. M. (1974) Rapid separation of panic ulate components and soluble cytoplasm of isolated rat-liver cells. Biochim. Biophys. Acta 333: 393-399.

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Biophys. Acta 268: 364-372. Allred, J. B. & Guy, D. G. (1969) Determination of coenzyme A and acetyl CoA in tissue extracts. Anal. Biochem. 29: 293-299. Berry, M. N. & Friend, D. S. (1969) High-yield preparation of isolated rat liver parenchymal cells. A biochemical and fine structural study. J. Cell Biol. 43: 506-520. Bonnett, R., Cannon, J. R., Clark, V. M., Johnson, A. W., Parker, L. R., Smith, L. E. & Todd, A. (1957) Chemistry of the vitamin B12 group. Part V. The structure of the chromophoric grouping. J. Chem. Soc. 1158-1168. Brass, E. P. J1992] Interaction of carnitine and propionate with pyruvate oxidation by hepatocytes from clofibrate-treated rats: importance of coenzyme A availability. J. Nutr. 122: 234—240. Brass, E. P. & Beyerinck, R. A. (1988) Effects of propionate and carnitine on the hepatic oxidation of short- and medium-chainlength fatty acids. Biochem. J. 250: 819-825. Brass, E. P. &. Ruff, L. J. (1991| Effect of clofibrate treatment on hepatic prostaglandin catabolism and action. J. Pharmacol. Exp. Ther. 257: 1034-1038. Brass, E. P., Tahiliani, A. G., Allen, R. H. & Stabler, S. P. (1990a) Coenzyme A metabolism in vitamin B-12-deficient rats. J. Nutr. 120: 290-297. Brass, E. P., Allen, R. H., Ruff, L. J. & Stabler, S. P. (1990b| Effect of hydroxycobalamin(c-lactam) on propionate and carnitine metabolism in the rat. Biochem. J. 266: 809-815. Bremer, J., Wojtczak, A. & Skrede, S. (1972) The leakage and destruction of CoA in isolated mitochondria. Eur. f. Biochem. 25: 190-197. Chalmers, R. A., Roe, C. R., Tracey, B. M., Stacey, T. E., Hoppel, C. L. & Millington, D. S. (1983) Secondary carnitine insufficiency in disorders of organic acid metabolism: modulation of acylCoA/CoASH ratios by L-carnitine. Biochem. Soc. Trans. 11: 724-725. DeDuve, C., Pressman, B. C., Gianetto, R., Wattiaux, R. & Appelmans, F. (1955) In tracellular distribution patterns of enzymes in rat-liver tissue. Biochem. J. 60: 604-617. Dolphin, P. (1971) Preparation of the reduced forms of vitamin 812 and of some analogs of the vitamin 812 coenzyme containing a cobalt-carbon bond. Methods Enzymol. 18: 34-52. Fenton, W. A., Hack, A. M., Helfgott, D. & Rosenberg, L. E. (1984) Biogenesis of the mitochondrial enzyme methylmalonyl-CoA mutase. J. Biol. Chem. 259: 6616-6621. Fisher, M. N. & Neely, J. R. (1985) Regulation of pantothenate kinase from various tissues of the rat. FEES Lett. 190: 293-296. Cornali, A. G., Bardawill, G. J. & David, M. M. (1949) Determi nation of serum proteins by means of the biuret reaction. J. Biol. Chem. 177: 751-766. Halvorsen, O. & Skrede, S. (1982) Regulation of the biosynthesis of CoA at the level of pantothenate kinase. Eur. J. Biochem. 124: 211-215. Hoppel, C., DiMarco, J. & Tandler, B. (1979) Riboflavin and rat hepatic cell structure and function. J. Biol. Chem. 254: 4164-4170. Ito, M., Ikeda, Y., Arnez, J. G., Finocchiaro, G. & Tanaka, K. (1990) The enzymatic basis for the metabolism and inhibitory effects of valproic acid: dehydrogenation of valproyl-CoA by 2-methylbranched-chain acyl-CoA dehydrogenase. Biochim. Biophys. Acta 1034: 213-218. Krahenbuhl, S., Ray, D. B., Stabler, S. P. & Brass, E. P. (1990) Increased hepatic mitochondrial capacity in rats with hydroxycobalamin[c-lactam]-induced methylmalonic aciduria. J. Clin. Invest. 86: 2054-2061. Lopaschuk, G. D. & Neely, J. R. (1987) Stimulation of myocardial

Rat hepatic coenzyme A is redistributed in response to mitochondrial acyl-coenzyme A accumulation.

Coenzyme A without an acyl-thioester (CoASH) is required for numerous cellular reactions, and sequestration of CoASH as acyl-CoAs may impair metabolic...
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