Abstract
cDNA clones corresponding to the entire length of mRNA for the alpha subunit of human pyruvate dehydrogenase (EC 1.2.4.1), the E1 component of the pyruvate dehydrogenase complex, have been isolated from liver cDNA libraries. Two classes of cDNA clones were obtained and these correspond to two forms of pyruvate dehydrogenase E1 alpha mRNA. Both mRNA species have been demonstrated in a variety of human tissues and cultured fibroblasts. The cDNA sequence has been determined and, from it, the protein sequence of the human E1 alpha subunit was deduced. The protein is synthesized with a typical mitochondrial import leader sequence and the peptide bond at which this sequence is cleaved after transport into the mitochondrion has been determined by direct amino acid sequencing of the mature E1 alpha subunit. The human pyruvate dehydrogenase E1 alpha subunit contains identical phosphorylation sites to those found in the corresponding porcine protein. Preliminary studies of pyruvate dehydrogenase E1 alpha mRNA in cultured fibroblasts from patients with severe pyruvate dehydrogenase deficiency have revealed considerable heterogeneity as would be expected from protein studies.
Highlights
ISOLATION OF cDNA CLONESFOR T H E Ela SUBUNIT,SEQUENCE ANALYSIS, AND CHARACTERIZATION OF T H E mRNA*
From the Murdoch Institutefor Research into BirthDefects, Royal ChildrenS Hospital and the $Departmentof Paediatrics, University of Melbourrze,Australia cDNA clones corresponding to the entire length of mRNA for the a subunit of human pyruvate dehydrogenase (EC 1.2.4.1), the E l component of the pyruvate dehydrogenase complex, have been isolated from liver cDNA libraries
Two classesof cDNA clones wereobtained and these correspond to two forms of pyruvate dehydrogenase Elm mRNA
Summary
ISOLATION OF cDNA CLONESFOR T H E Ela SUBUNIT,SEQUENCE ANALYSIS, AND CHARACTERIZATION OF T H E mRNA*. TheEla subunit of the pyruvate dehydrogenase complex is of particular interest, as in addition to its role in the E l enzyme reaction, it contains the phosphorylation sites which regulate the activityof the whole complex. In man, this subunit is of specific importance as iits the siteof the genetic defect in the majority of patients with pyruvatedehydrogenase deficiency [4]. We present thecDNA sequence corresponding to the complete mRNA, the predicted amino acid sequence of the precursor, and mature protein and preliminary resuolftsstudies of the pyruvate dehydrogenase Ela gene and mRNA in patients with genetic defects in this subunit.
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