The Aromatization of Cyclohexanecarboxyl-CoA to Hippuric Acid by Guinea Pig Liver Mitochondria: Submitochondrial Localization

Overview

Journal Mol Cell Biochem

Publisher Springer

Specialty Biochemistry

Date 1985 Jul 1

PMID 4047029

Citations 1

Authors

A M Svardal

R R SCHELINE

Affiliations

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Abstract

The conversion of cyclohexanecarboxyl-CoA to hippuric acid in submitochondrial fractions from guinea pig liver was studied using a gas chromatographic-mass spectrometric method employing selected ion monitoring. Comparison of the activities of the cyclohexanecarboxyl-CoA to hippuric acid converting system (CCoAHC-system) and marker enzymes in the various submitochondrial fractions showed that the CCoAHC-system is localized in the mitochondrial matrix. Partial separation of the inner and outer membranes has been accomplished by treating mitochondria with digitonin in isotonic medium and fractionating the treated mitochondria by differential centrifugation. A digitonin-protein ratio of 2.6 mg of digitonin/10 mg of protein must be used in order to release significant amounts of amine oxidase activity (outer membrane marker) from low speed mitochondrial pellets. This pellet still contained most of the glutamate dehydrogenase activity and was insignificantly contaminated with adenylate kinase. Moderate concentrations of phenazine methosulfate (PMS) greatly stimulated the activity of the CCoAHC-system, even in intact mitochondria (optimal concentration of PMS: 1 mM) whilst higher concentrations (greater than 1 mM) decreased the activity. The formation of hippuric acid in these mitochondrial preparations was linear with time for at least 40 min and linear with respect to protein concentration up to approximately 2.0 mg mitochondrial protein X ml-1.

Citing Articles

The aromatization of cyclohexanecarboxylic acid to hippuric acid: substrate specificity and species differences.

Svardal A, SCHELINE R Mol Cell Biochem. 1985; 67(2):171-9.

PMID: 4047028 DOI: 10.1007/BF02370176.

References

Babior B, Bloch K . Aromatization of cyclohexanecarboxylic acid. J Biol Chem. 1966; 241(16):3643-51. View

Mitoma C, POSNER H, Leonard F . Aromatization of hexahydrobenzoic acid by mamalian liver mitochondria. Biochim Biophys Acta. 1958; 27(1):156-60. DOI: 10.1016/0006-3002(58)90302-0. View

Hall C . Acyl-CoA dehydrogenases from pig liver mitochondria. Methods Enzymol. 1981; 71 Pt C:375-85. DOI: 10.1016/0076-6879(81)71047-4. View

Husain M, Steenkamp D . Electron transfer flavoprotein from pig liver mitochondria. A simple purification and re-evaluation of some of the molecular properties. Biochem J. 1983; 209(2):541-5. PMC: 1154123. DOI: 10.1042/bj2090541. View

Groot P . Acyl-CoA synthetases in guinea-pig liver mitochondria. Purification and characterization of a distinct propionyl-CoA synthetase. Biochim Biophys Acta. 1976; 441(2):260-7. DOI: 10.1016/0005-2760(76)90169-7. View

SINGER T . Determination of the activity of succinate, NADH, choline, and alpha-glycerophosphate dehydrogenases. Methods Biochem Anal. 1974; 22:123-75. DOI: 10.1002/9780470110423.ch3. View

James M, Bend J . A radiochemical assay for glycine N-acyltransferase activity. Some properties of the enzyme in rat and rabbit. Biochem J. 1978; 172(2):285-91. PMC: 1185694. DOI: 10.1042/bj1720285. View

Goldman P, VAGELOS P . The specificity of triglyceride synthesis from diglycerides in chicken adipose tissue. J Biol Chem. 1961; 236:2620-3. View

Ikeda Y, Dabrowski C, Tanaka K . Separation and properties of five distinct acyl-CoA dehydrogenases from rat liver mitochondria. Identification of a new 2-methyl branched chain acyl-CoA dehydrogenase. J Biol Chem. 1983; 258(2):1066-76. View

10.

Svardal A, SCHELINE R . Intracellular localization and some properties of the system in guinea pig liver responsible for the aromatization of cyclohexanecarboxylic acid to hippuric acid. Mol Cell Biochem. 1985; 65(2):107-15. DOI: 10.1007/BF00221093. View

11.

Szasz G, Gerhardt W, Gruber W, BERNT E . Creatine kinase in serum: 2. Interference of adenylate kinase with the assay. Clin Chem. 1976; 22(11):1806-11. View

12.

Haddock B, Yates D, Garland P . The localization of some coenzyme A-dependent enzymes in rat liver mitochondria. Biochem J. 1970; 119(3):565-73. PMC: 1179388. DOI: 10.1042/bj1190565. View

13.

Schnaitman C, Erwin V, Greenawalt J . The submitochondrial localization of monoamine oxidase. An enzymatic marker for the outer membrane of rat liver mitochondria. J Cell Biol. 1967; 32(3):719-35. PMC: 2107278. DOI: 10.1083/jcb.32.3.719. View

14.

Aas M . Organ and subcellular distribution of fatty acid activating enzymes in the rat. Biochim Biophys Acta. 1971; 231(1):32-47. DOI: 10.1016/0005-2760(71)90253-0. View

15.

Arrigoni O, SINGER T . Limitations of the phenazine methosulphate assay for succinic and related dehydrogenases. Nature. 1962; 193:1256-8. DOI: 10.1038/1931256a0. View

16.

Gatley S, Sherratt H . The synthesis of hippurate from benzoate and glycine by rat liver mitochondria. Submitochondrial localization and kinetics. Biochem J. 1977; 166(1):39-47. PMC: 1164954. DOI: 10.1042/bj1660039. View

17.

Bendall D, DE DUVE C . Tissue-fractionation studies. 14. The activation of latent dehydrogenases in mitochondria from rat liver. Biochem J. 1960; 74:444-50. PMC: 1204240. DOI: 10.1042/bj0740444. View