Complete Defect of the E1,f Subunit of the Branched Chain a-Ketoacid Dehydrogenase Complex Due to a Deletion of an 11-bp Repeat Sequence which Encodes a Mitochondrial Targeting Leader Peptide in a Family with the Disease

Branched chain a-ketoacid dehydrogenase (BCKDH) deficiency results in maple syrup urine disease(MSUD).Weexamined the molecular basis of familial cases ofMSUD by analyzing the activity, subunit structure, mRNA sequence, and genome structure ofthe affected enzyme. TheBCKDH activity in the proband withMSUD was 6% ofthe normal control level. Immunoblot analysis revealed that the ElB subunit ofBCKDH was absent and that the Ela subunit ofBCKDH was markedly reduced. We amplified the cDNAs of the Ela subunit and the Elf subunit of theBCKDH complex obtained from cells of the patient, using the polymerase chain reaction method, then sequenced the amplified cDNAs. The deduced amino acid sequence for the Ela subunit of the patient's cell was normal. An 11-bp deletion was identified in the region that encoded the mitochondrial targeting leader peptide in the Elft cDNA. This 11-bp sequence is found in the first exon of the BCKDH-Elf# gene, as a direct tandem repeat. Amplification ofgenomicDNA revealed that the consanguineous parents were heterozygous for this mutant allele, and sister and brother ofthe patient with the disease were homozygous for this mutant allele. This 11-bp deletion mutation caused a change in the reading frame and the mature El: protein was defective. These observations show the biological importance of the El,@ subunit ofBCKDH to maintain normal function of the enzyme activity. The absence ofthe ElfI subunit results in instability of the Ela subunit. (J. Clin.