[Gene analysis of maple syrup urine disease (MSUD)]

Maple syrup urine disease (MSUD), an autosomal recessive hereditary metabolic disorder, is due to defective oxidative decarboxylation of the branched-chain alpha-ketoacids (BCKAs) derived from transamination of the three branched-chain amino acids, valine, leucine and isoleucine. The oxidative decarboxylation of three BCKAs is catalysed by the branched-chain alpha-ketoacid dehydrogenase (BCKDH) complex. BCKDH consists of three catalytic components: E1, E2 and E3. The E1 component is further composed of two subunits, E1 alpha and E1 beta. To clarify the mechanisms involved in MSUD, measurements of the enzyme activity in cultured cells, measurements of the generation time in cultured cells, complementation analysis and immunoblot analysis were performed. To further elucidate the molecular mechanisms of MSUD, we and others isolated and characterized cDNAs encoding BCKDH-E1 alpha, E1 beta, E2 and E3. The human genome structures of BCKDH -E1 alpha, E1 beta and E2 were also characterized. Gene mutations in E1 alpha, E1 beta and E2, respectively, were identified at the molecular level in three cases of classical MSUD. It became clear that the molecular mechanisms of MSUD involved not only the function of each subunit but also the protein-protein interactions between each subunit. In an attempt to further analyse the molecular basis of MSUD, we carried out complementation analyses by somatic cell hybridization, and identified the affected component of BCKDH complex in the MSUD patient. Furthermore, to rapidly screen for gene mutations, we used PCR-SSCP analysis. Seventeen patients with MSUD were examined using these methods. Defects of E1 alpha, E1 beta and E2 subunits were suspected in 8, 5, and 4 patients, respectively, by complementation analysis.(ABSTRACT TRUNCATED AT 250 WORDS)