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. 1990 Sep;86(3):1000–1003. doi: 10.1172/JCI114761

Molecular basis of medium chain acyl-coenzyme A dehydrogenase deficiency. An A to G transition at position 985 that causes a lysine-304 to glutamate substitution in the mature protein is the single prevalent mutation.

I Yokota 1, Y Indo 1, P M Coates 1, K Tanaka 1
PMCID: PMC296821  PMID: 2394825

Abstract

We sequenced polymerase chain reaction (PCR)-amplified variant medium chain acyl-CoA dehydrogenase (MCAD) cDNAs in cultured fibroblasts from three MCAD-deficient patients. In all three patients, an A to G transition was identified at position 985 of the coding region. Since no appropriate restriction sites for detecting this point mutation were found, we devised a PCR method that amplifies an 87-bp fragment from position 955. In the 5' primer encompassing positions 955 to 984, A-981 was artificially substituted with C. With the presence of C-981 and G-985, an Nco I restriction site is introduced in the mutant copies. When cDNA or genomic DNA from fibroblasts of nine MCAD-deficient patients were tested with this method, the copies from all of them completely cleaved into two shorter fragments by Nco I, indicating their homozygosity for the A----G-985 transition. In contrast, the copies from all eight controls remained intact. Thus, this A----G-985 transition is the single prevalent mutation causing MCAD deficiency, a highly unusual feature for any genetic disorder. The PCR/Nco I digestion method is suitable for the diagnosis of MCAD deficiency.

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Selected References

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  1. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Duran M., Hofkamp M., Rhead W. J., Saudubray J. M., Wadman S. K. Sudden child death and 'healthy' affected family members with medium-chain acyl-coenzyme A dehydrogenase deficiency. Pediatrics. 1986 Dec;78(6):1052–1057. [PubMed] [Google Scholar]
  3. Ikeda Y., Hale D. E., Keese S. M., Coates P. M., Tanaka K. Biosynthesis of variant medium chain acyl-CoA dehydrogenase in cultured fibroblasts from patients with medium chain acyl-CoA dehydrogenase deficiency. Pediatr Res. 1986 Sep;20(9):843–847. doi: 10.1203/00006450-198609000-00007. [DOI] [PubMed] [Google Scholar]
  4. Ikeda Y., Keese S. M., Fenton W. A., Tanaka K. Biosynthesis of four rat liver mitochondrial acyl-CoA dehydrogenases: in vitro synthesis, import into mitochondria, and processing of their precursors in a cell-free system and in cultured cells. Arch Biochem Biophys. 1987 Feb 1;252(2):662–674. doi: 10.1016/0003-9861(87)90072-5. [DOI] [PubMed] [Google Scholar]
  5. Ikeda Y., Keese S. M., Tanaka K. Molecular heterogeneity of variant isovaleryl-CoA dehydrogenase from cultured isovaleric acidemia fibroblasts. Proc Natl Acad Sci U S A. 1985 Oct;82(20):7081–7085. doi: 10.1073/pnas.82.20.7081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Ikeda Y., Okamura-Ikeda K., Tanaka K. Purification and characterization of short-chain, medium-chain, and long-chain acyl-CoA dehydrogenases from rat liver mitochondria. Isolation of the holo- and apoenzymes and conversion of the apoenzyme to the holoenzyme. J Biol Chem. 1985 Jan 25;260(2):1311–1325. [PubMed] [Google Scholar]
  7. Kelly D. P., Kim J. J., Billadello J. J., Hainline B. E., Chu T. W., Strauss A. W. Nucleotide sequence of medium-chain acyl-CoA dehydrogenase mRNA and its expression in enzyme-deficient human tissue. Proc Natl Acad Sci U S A. 1987 Jun;84(12):4068–4072. doi: 10.1073/pnas.84.12.4068. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kim J. J., Wu J. Structure of the medium-chain acyl-CoA dehydrogenase from pig liver mitochondria at 3-A resolution. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6677–6681. doi: 10.1073/pnas.85.18.6677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kølvraa S., Gregersen N., Christensen E., Hobolth N. In vitro fibroblast studies in a patient with C6-C10-dicarboxylic aciduria: evidence for a defect in general acyl-CoA dehydrogenase. Clin Chim Acta. 1982 Nov 24;126(1):53–67. doi: 10.1016/0009-8981(82)90361-8. [DOI] [PubMed] [Google Scholar]
  10. Matsubara Y., Indo Y., Naito E., Ozasa H., Glassberg R., Vockley J., Ikeda Y., Kraus J., Tanaka K. Molecular cloning and nucleotide sequence of cDNAs encoding the precursors of rat long chain acyl-coenzyme A, short chain acyl-coenzyme A, and isovaleryl-coenzyme A dehydrogenases. Sequence homology of four enzymes of the acyl-CoA dehydrogenase family. J Biol Chem. 1989 Sep 25;264(27):16321–16331. [PubMed] [Google Scholar]
  11. Matsubara Y., Kraus J. P., Ozasa H., Glassberg R., Finocchiaro G., Ikeda Y., Mole J., Rosenberg L. E., Tanaka K. Molecular cloning and nucleotide sequence of cDNA encoding the entire precursor of rat liver medium chain acyl coenzyme A dehydrogenase. J Biol Chem. 1987 Jul 25;262(21):10104–10108. [PubMed] [Google Scholar]
  12. McCabe E. R., Huang S. Z., Seltzer W. K., Law M. L. DNA microextraction from dried blood spots on filter paper blotters: potential applications to newborn screening. Hum Genet. 1987 Mar;75(3):213–216. doi: 10.1007/BF00281061. [DOI] [PubMed] [Google Scholar]
  13. Naito E., Indo Y., Tanaka K. Identification of two variant short chain acyl-coenzyme A dehydrogenase alleles, each containing a different point mutation in a patient with short chain acyl-coenzyme A dehydrogenase deficiency. J Clin Invest. 1990 May;85(5):1575–1582. doi: 10.1172/JCI114607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Naito E., Indo Y., Tanaka K. Short chain acyl-coenzyme A dehydrogenase (SCAD) deficiency. Immunochemical demonstration of molecular heterogeneity due to variant SCAD with differing stability. J Clin Invest. 1989 Nov;84(5):1671–1674. doi: 10.1172/JCI114346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Naito E., Ozasa H., Ikeda Y., Tanaka K. Molecular cloning and nucleotide sequence of complementary DNAs encoding human short chain acyl-coenzyme A dehydrogenase and the study of the molecular basis of human short chain acyl-coenzyme A dehydrogenase deficiency. J Clin Invest. 1989 May;83(5):1605–1613. doi: 10.1172/JCI114058. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Rinaldo P., O'Shea J. J., Coates P. M., Hale D. E., Stanley C. A., Tanaka K. Medium-chain acyl-CoA dehydrogenase deficiency. Diagnosis by stable-isotope dilution measurement of urinary n-hexanoylglycine and 3-phenylpropionylglycine. N Engl J Med. 1988 Nov 17;319(20):1308–1313. doi: 10.1056/NEJM198811173192003. [DOI] [PubMed] [Google Scholar]
  17. 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]

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