BMB Reports

Korean Society for Biochemistry and Molecular Biology (생화학분자생물학회)
권호 표지
DOI QR코드

DOI QR Code

Rat Malonyl-CoA Decarboxylase; Cloning, Expression in E. coli and its Biochemical Characterization

  • Lee, Gha-Young (Department of Biochemistry, College of Science, Protein Network Research Center, Yonsei University) ;
  • Bahk, Young-Yil (Department of Biochemistry, College of Science, Protein Network Research Center, Yonsei University) ;
  • Kim, Yu-Sam (Department of Biochemistry, College of Science, Protein Network Research Center, Yonsei University)
  • Published : 2002.03.31
Download PDF
⟨ Previous Next ⟩

Abstract

Malonyl-CoA decarboxylase (E.C.4.1.1.9) catalyzes the conversion of malonyl-CoA to acetyl-CoA. Although the metabolic role of this enzyme has not been fully defined, it has been reported that its deficiency is associated with mild mental retardation, seizures, hypotonia, cadiomyopathy, developmental delay, vomiting, hypoglycemia, metabolic acidosis, and malonic aciduria. Here, we isolated a cDNA clone for malonyl CoA decarboxylase from a rat brain cDNA library, expressed it in E. coli, and characterized its biochemical properties. The full-length cDNA contained a single open-reading frame that encoded 491 amino acid residues with a calculated molecular weight of 54, 762 Da. Its deduced amino acid sequence revealed a 65.6% identity to that from the goose uropigial gland. The sequence of the first 38 amino acids represents a putative mitochondrial targeting sequence, and the last 3 amino acid sequences (SKL) represent peroxisomal targeting ones. The expression of malonyl CoA decarboxylase was observed over a wide range of tissues as a single transcript of 2.0 kb in size. The recombinant protein that was expressed in E. coli was used to characterize the biochemical properties, which showed a typical Michaelis-Menten substrate saturation pattern. The $K_m$ and $V_{max}$ were calculated to be $68\;{\mu}M$ and $42.6\;{\mu}mol/min/mg$, respectively.

Keywords

References

  1. An, J. H. and Kim, Y. S. (1998) A novel gene cluster encoding malonyl-CoA decarboxylase (MatA), malonyl-CoA synthetase (MatB) and a putative dicarboxylate carrier protein (MatC) in Rhizobium trifolii: cloning, sequencing, and expression of the enzymes in Escherichia coli. Eur. J. Biochem. 257, 395-402. https://doi.org/10.1046/j.1432-1327.1998.2570395.x
  2. An, J. H., Lee G. Y., Song, J. H, Lee, D. W. and Kim, Y. S. (1999) Properties of malonyl-CoA decarboxylase from Rhizobium trifolii. J. Biochem. Mol. BioI. 32, 414-418.
  3. Babior, B. M. (1973) Cleavage of coenzyme B12 by methylmalonyl Coenzyme A mutase. J. BioI. Chem. 248, 1445-1450.
  4. Byun, H. S. and Kim, Y. S. (1995) CoA transferase and malonyl-CoA decarboxylase activity of malonate decarboxylase from Acinotobacter calcoaceticus. J. Biochem. Mol. BioI. 28, 107-111.
  5. Courchesne-Smith, C., Jang, S. H., Shi, Q., DeWille, J., Sasaki, G. and Kolattukudy, P. E. (1992) Cytoplasmic accumulation of a normally mitochondrial malonyl-CoA decarboxylase by the use of an alternate transcription start site. Arch. Biochem. Biophys. 298, 576-586. https://doi.org/10.1016/0003-9861(92)90452-3
  6. Dickson, A. C., McEvoy, J. A. and Koeppen, A. H. (1994) The cellular localization of malonyl-coenzyme A decarboxylase in rat brain. Neurochem. Res. 19, 1271-1276. https://doi.org/10.1007/BF01006817
  7. Eun, S. H., Kim, D. J. and Kim, Y. S. (2001) Acinetobacter calcoaceticus Glucose-1-phosphate thyrnidytransferase; Cloning, sequencing, and expression in E. coli. J. Biochem. Mol. Biol. 34, 230-236.
  8. Gregg, A. R., Warman, A. W., Thorbum, D. R. and O'Brien, W. E. (1998) Combined malonic and methylmalonic aciduria with normal malonyl-coenzyme A decarboxylase activity: a case supporting multiple aetiologies. J. Inherit. Metab. Dis. 21, 382-390. https://doi.org/10.1023/A:1005302607897
  9. Haan, E. A., Scholem, R. D., Croll, H. B. and Brown, G. K. (1986) Malonyl coenzyme A decarboxylase deficiency. Clinical and biochemical findings in a second child with a more severe enzyme defect. Eur. J. Pediatr. 144, 567-570. https://doi.org/10.1007/BF00496037
  10. Jang, S. H., Cheesbrough, T. M. and Kolattukudy, P. E. (1989) Molecular cloning, nucleotide sequence and tissue distribution of malonyl-CoA decarboxylase. J. Biol. Chem. 264, 3500-3505.
  11. Kaushik, V. K., Young, M. E., Dean, D. J., Kurowski, T. G., Saha， A. K. and Ruderman, N. B. (2001) Regulation of fatty acid oxidation and glucose metabolism in rat soleus muscle: effects of AICAR. Am. J. Physiol. Endocrinol. Metab. 281, E335- E340.
  12. Kim, Y. S. and Kolattukudy, P. E. (1978a) Purification and properties of malonyl-CoA decarboxylase from rat liver rnitochondria and its immunological comparison with the enzymes from rat brain, heart, and man1ffi없y gland. Arch. Biochem. Biophys. 190, 224-246.
  13. Kim, Y. S. and Kolattukudy, P. E. (1978b) Malonyl-CoA decarboxylase from the uropygial gland of waterfowl: purification, properties, immunological comparison, and role in regulating the synthesis of multimethyl-branched fatty acids. Arch. Biochem Biophys. 190, 585-597. https://doi.org/10.1016/0003-9861(78)90314-4
  14. Kolattukudy, P. E., Poulose, A. J. and Kim, Y. S. (1981) Malonyl-CoA decarboxylase from avian, mammalian, and rnicrobial sources. Methods Enzymol. 71C, 150-153.
  15. Kozak, M. (1996) Interpreting cDNA sequences: some insights from studies on translation. Mamm. Genome 7, 563-574 https://doi.org/10.1007/s003359900171
  16. Krawinkel, M. B., Oldigs, H. D., Santer, R., Lehnert, W., Wendel, U. and Schaub, J. (1994) Association of malonyl-CoA decarboxylase deficiency and heterozygote state for hemoglobin C disease. J. lnherit. Metab. Dis. 17, 636-637. https://doi.org/10.1007/BF00711609
  17. Lee, H. Y. and Kim, Y. S. (2001) Identification of amino acid residues in the carboxyl terminus required for malonate-responsive transcriptional regulation of MatR in Rhizobium leguminosarum bv. trifolii. J. Biochem Mol. Biol. 34, 305-309.
  18. McGarry, J. D. and Brown, N. F. (1997) The rnitochondrial carnitine pa1rnitoyltransferase system. From concept to molecular analysis. Eur. J. Biochem. 244, 1-14. https://doi.org/10.1111/j.1432-1033.1997.00001.x
  19. Park, S. J. and Cho, Y. D. (2000) Studies on the active site of the Arabidopsis thaliana S-Adenosylmethionine Decarboxy: Lys81 residue involvement in catalytic activity. J. Biachem. Mol. Biol. 33, 69-74.
  20. Prentki, M (1996) New insights into pancreatic beta-cell metabolic signaling in insulin secretion. Eur. J. Endocrinol. 134, 272-286. https://doi.org/10.1530/eje.0.1340272
  21. Ozand, P. T., Nyhan, W. L., al Aqeel, A. and Christodoulou, J. (1994) Malonic aciduria. Brain. Dev. 16, 7-1 1. https://doi.org/10.1016/0387-7604(94)90091-4
  22. Sacksteder, K. A., Morrell, J. C., Wanders, R. J., Matalon, R. and Gould, S. J. (1999) MCD encodes peroxisomal and cytoplasmic forms of malonyl-CoA decarboxylase and is mutated in malonyl-CoA decarboxylase deficiency. J. Biol. Chem. 274, 24461-24468. https://doi.org/10.1074/jbc.274.35.24461
  23. Sarnbrook, J., Fritsch, E. F. and Maniatis, T. (1989) Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
  24. Scrutton, M. C. and Utter, M. F. (1967) Pyruvate carboxylase IX, some properties of the activation by certain acyl derivatives of coenzyme A. J. Biol. Chem. 242, 1723-1735.
  25. Sha, Z., Stabel, T. J. and Mayfield, J. E. (1994) Brucella abortus cata1ase is a periplasrnic protein lacking a standard signal sequence. J. Bacterial. 176, 7375-7377. https://doi.org/10.1128/jb.176.23.7375-7377.1994
  26. Yano, S., Sweetman, L., Thorbum, D. R., Mofidi, S. and Williams, J. C. (1997) A new case of malonyl coenzyme A decarboxylase deficiency presenting with cardiomyopathy. Eur. J. Pediatr. 156, 382-383 https://doi.org/10.1007/s004310050619

Cited by

  1. Malonyl-CoA decarboxylase (MCD) is differentially regulated in subcellular compartments by 5′AMP-activated protein kinase (AMPK) vol.271, pp.13, 2004, https://doi.org/10.1111/j.1432-1033.2004.04218.x
  2. Isolation of two distinct populations of recombinant antibody molecules specific for rat malonyl-CoA decarboxylase from a semi-synthetic human scFv display library using Ex-phage system vol.91, pp.2-3, 2004, https://doi.org/10.1016/j.imlet.2003.11.010
  3. Genomic organization and characterization of the promoter of rat malonyl-CoA decarboxylase gene vol.1577, pp.1, 2002, https://doi.org/10.1016/S0167-4781(02)00398-6
  4. Expression, purification, and characterization of human malonyl-CoA decarboxylase vol.34, pp.2, 2004, https://doi.org/10.1016/j.pep.2003.11.023
  5. Assay of the activity of malonyl–coenzyme A decarboxylase by gas chromatography–mass spectrometry vol.363, pp.2, 2007, https://doi.org/10.1016/j.ab.2007.01.010
  6. Crystal Structures of Malonyl-Coenzyme A Decarboxylase Provide Insights into Its Catalytic Mechanism and Disease-Causing Mutations vol.21, pp.7, 2013, https://doi.org/10.1016/j.str.2013.05.001
  7. A highly sensitive high-throughput luminescence assay for malonyl-CoA decarboxylase vol.376, pp.1, 2008, https://doi.org/10.1016/j.ab.2008.01.033