title: Mechanisms of expression of pyruvate dehydrogenase deficiency caused by an E1alpha subunit mutation.
abstract: OBJECTIVE
  To characterize the biochemical mechanisms of expression of the pyruvate dehydrogenase (PDH) E1alpha subunit exon 10 R302C missense mutation.
  
  
  BACKGROUND
  Mutations in the X-linked E1alpha subunit gene are responsible for most cases of PDH deficiency, an important cause of neurodevelopmental defects and neurodegeneration with primary lactic acidemia. Although the disease shows extreme allelic heterogeneity, the R302C mutation has been defined in several unrelated cases.
  
  
  METHODS
  Cell lines expressing selectively either the mutant or wild-type E1alpha alleles against identical genetic backgrounds were generated from the fibroblasts of a female heterozygous for the R302C mutation. Enzyme activity, mRNA, polypeptide expression, and turnover were studied in each.
  
  
  RESULTS
  The residual PDH activity was below measurable levels in the cell line (B5) expressing only the mutant allele and normal in the wild-type polypeptide expressing (A10) cell line, confirming that the R302C mutation alone is sufficient to cause a severe PDH deficiency. The mutant polypeptide was less stable than the wild-type polypeptide, but the steady-state level of the mutant E1alpha protein was reduced only two- to threefold.
  
  
  CONCLUSIONS
  The primary mechanism of expression of the R302C mutation must be limitation of catalytic efficiency. We speculate that catalysis may be inhibited in the mutant polypeptide because conformational changes are induced near serine 300, a residue that is particularly important as a regulatory phosphorylation site in the wild-type polypeptide.
authors.affiliation: Institute of Molecular Medicine, Department of Clinical Neurology, University of Oxford, UK.
authors.forename: K J
authors.initials: KJ
authors.lastname: Morten
authors.forename: M
authors.initials: M
authors.lastname: Caky
authors.forename: P M
authors.initials: PM
authors.lastname: Matthews
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D002478:Cells, Cultured; D002648:Child; D005260:Female; D005347:Fibroblasts / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic; D006580:Genetic Carrier Screening; D006801:Humans; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics*; D015325:Pyruvate Dehydrogenase Complex Deficiency Disease / Q000201:enzymology / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis; D012867:Skin / Q000201:enzymology; D014158:Transcription, Genetic; D014960:X Chromosome
chemical_list: D011768:Pyruvate Dehydrogenase Complex; D012333:RNA, Messenger