Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Sep 15;326(Pt 3):883–889. doi: 10.1042/bj3260883

Molecular cloning of cDNA species for rat and mouse liver alpha-methylacyl-CoA racemases.

W Schmitz 1, H M Helander 1, J K Hiltunen 1, E Conzelmann 1
PMCID: PMC1218746  PMID: 9307041

Abstract

cDNA species coding for alpha-methylacyl-CoA racemase were cloned from rat and mouse liver cDNA libraries and characterized. The rat liver lambdagt11 cDNA expression library was screened with anti-racemase IgG [Schmitz, Albers, Fingerhut and Conzelmann (1995) Eur. J. Biochem.231, 815-822]. Several full-length clones were obtained that contained an open reading frame of 1083 bp, coding for a protein of 361 amino acid residues with a predicted molecular mass of 39679 Da. The sequences of three peptides that were isolated by HPLC from a tryptic digest of purified rat liver racemase fully matched the cDNA-derived amino acid sequence. The cDNA coding for mouse racemase was cloned from a mouse liver lambdaZAP cDNA expression library and sequenced. The coding region of 1080 bp codes for a 360-residue protein (molecular mass 39558 Da) that shares 89.7% similarity with the rat protein. Expression of the rat racemase as are combinant protein in Escherichia coli with the pTrcHisB-expression vector yielded enzymically active protein. The amino acid sequences of alpha-methylacyl-CoA racemases do not resemble any known sequence of beta-oxidation or auxiliary enzymes, supporting the view of a highly diverse evolutionary origin of enzymes acting on fatty acyl-CoA S-esters.

Full Text

The Full Text of this article is available as a PDF (971.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ackman R. G., Hansen R. P. The occurrence of diastereomers of phytanic and pristanic acids and their determination by gas-liquid chromatography. Lipids. 1967 Sep;2(5):357–362. doi: 10.1007/BF02531848. [DOI] [PubMed] [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  3. Avigan J., Steinberg D., Gutman A., Mize C. E., Milne G. W. Alpha-decarboxylation, an important pathway for degradation of phytanic acid in animals. Biochem Biophys Res Commun. 1966 Sep 22;24(6):838–844. doi: 10.1016/0006-291x(66)90324-x. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Engel C., Wierenga R. The diverse world of coenzyme A binding proteins. Curr Opin Struct Biol. 1996 Dec;6(6):790–797. doi: 10.1016/s0959-440x(96)80009-1. [DOI] [PubMed] [Google Scholar]
  6. Fingerhut R., Schmitz W., Conzelmann E. Accumulation of phytanic acid alpha-oxidation intermediates in Zellweger fibroblasts. J Inherit Metab Dis. 1993;16(3):591–594. doi: 10.1007/BF00711690. [DOI] [PubMed] [Google Scholar]
  7. Hansen R. P., Morrison J. D. The isolation and identification of 2,6,10,14-tetramethylpentadecanoic acid from butterfat. Biochem J. 1964 Nov;93(2):225–228. doi: 10.1042/bj0930225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Pedersen J. I., Gustafsson J. Conversion of 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestanoic acid into cholic acid by rat liver peroxisomes. FEBS Lett. 1980 Dec 1;121(2):345–348. doi: 10.1016/0014-5793(80)80377-2. [DOI] [PubMed] [Google Scholar]
  9. Pedersen J. I., Veggan T., Björkhem I. Substrate stereospecificity in oxidation of (25S)-3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestanoyl-CoA by peroxisomal trihydroxy-5 beta-cholestanoyl-CoA oxidase. Biochem Biophys Res Commun. 1996 Jul 5;224(1):37–42. doi: 10.1006/bbrc.1996.0981. [DOI] [PubMed] [Google Scholar]
  10. Poulos A., Sharp P., Fellenberg A. J., Johnson D. W. Accumulation of pristanic acid (2, 6, 10, 14 tetramethylpentadecanoic acid) in the plasma of patients with generalised peroxisomal dysfunction. Eur J Pediatr. 1988 Feb;147(2):143–147. doi: 10.1007/BF00442211. [DOI] [PubMed] [Google Scholar]
  11. Poulos A., Sharp P., Whiting M. Infantile Refsum's disease (phytanic acid storage disease): a variant of Zellweger's syndrome? Clin Genet. 1984 Dec;26(6):579–586. doi: 10.1111/j.1399-0004.1984.tb01107.x. [DOI] [PubMed] [Google Scholar]
  12. Purdue P. E., Lazarow P. B. Targeting of human catalase to peroxisomes is dependent upon a novel COOH-terminal peroxisomal targeting sequence. J Cell Biol. 1996 Aug;134(4):849–862. doi: 10.1083/jcb.134.4.849. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Reichel C., Brugger R., Bang H., Geisslinger G., Brune K. Molecular cloning and expression of a 2-arylpropionyl-coenzyme A epimerase: a key enzyme in the inversion metabolism of ibuprofen. Mol Pharmacol. 1997 Apr;51(4):576–582. doi: 10.1124/mol.51.4.576. [DOI] [PubMed] [Google Scholar]
  14. Roise D., Horvath S. J., Tomich J. M., Richards J. H., Schatz G. A chemically synthesized pre-sequence of an imported mitochondrial protein can form an amphiphilic helix and perturb natural and artificial phospholipid bilayers. EMBO J. 1986 Jun;5(6):1327–1334. doi: 10.1002/j.1460-2075.1986.tb04363.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Schmitz W., Albers C., Fingerhut R., Conzelmann E. Purification and characterization of an alpha-methylacyl-CoA racemase from human liver. Eur J Biochem. 1995 Aug 1;231(3):815–822. doi: 10.1111/j.1432-1033.1995.tb20766.x. [DOI] [PubMed] [Google Scholar]
  17. Schmitz W., Conzelmann E. Stereochemistry of peroxisomal and mitochondrial beta-oxidation of alpha-methylacyl-CoAs. Eur J Biochem. 1997 Mar 1;244(2):434–440. doi: 10.1111/j.1432-1033.1997.00434.x. [DOI] [PubMed] [Google Scholar]
  18. Schmitz W., Fingerhut R., Conzelmann E. Purification and properties of an alpha-methylacyl-CoA racemase from rat liver. Eur J Biochem. 1994 Jun 1;222(2):313–323. doi: 10.1111/j.1432-1033.1994.tb18870.x. [DOI] [PubMed] [Google Scholar]
  19. Shefer S., Cheng F. W., Batta A. K., Dayal B., Tint G. S., Salen G., Mosbach E. H. Stereospecific side chain hydroxylations in the biosynthesis of chenodeoxycholic acid. J Biol Chem. 1978 Sep 25;253(18):6386–6392. [PubMed] [Google Scholar]
  20. Van Veldhoven P. P., Croes K., Asselberghs S., Herdewijn P., Mannaerts G. P. Peroxisomal beta-oxidation of 2-methyl-branched acyl-CoA esters: stereospecific recognition of the 2S-methyl compounds by trihydroxycoprostanoyl-CoA oxidase and pristanoyl-CoA oxidase. FEBS Lett. 1996 Jun 10;388(1):80–84. doi: 10.1016/0014-5793(96)00508-x. [DOI] [PubMed] [Google Scholar]
  21. de Hoop M. J., Ab G. Import of proteins into peroxisomes and other microbodies. Biochem J. 1992 Sep 15;286(Pt 3):657–669. doi: 10.1042/bj2860657. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES