MOLECULAR CHARACTERIZATION OF PYRUVATE-CARBOXYLASE DEFICIENCY IN 2 CONSANGUINEOUS FAMILIES

Pyruvate carboxylase (PC) is a biotinylated mitochondrial enzyme that catalyzes the conversion of pyruvate to oxaloacetate. Children with in born errors of PC metabolism have lactic acidosis, hypoglycemia, and m ental retardation. The variable severity of the clinical phenotype is dependent on both genetic and environmental factors. Two consanguineou s families with moderate forms of PC deficiency were characterized at the biochemical and molecular levels. In both families, the probands w ere found to have low PC activity (range, 2-25% of control) in blood l ymphocytes and skin fibroblasts associated with either diminished or n ormal protein levels. In the first case, sequencing of patient-specifi c PC cDNA demonstrated a T to C substitution at nucleotide 434, which causes a valine to alanine change at amino acid residue 145. Direct se quencing of the parents showed that they are heterozygous for this mut ation. In the second family, a brother and sister had mental retardati on and episodes of severe lactic/ketoacidosis in early childhood. In t hese cases, a C to T substitution at nucleotide 1351 results in a cyst eine for arginine substitution at amino acid residue 451; the parents were also found to be heterozygous for this mutation. In both families , no other mutations were found, and both substitutions occurred in re latively conserved amino acid residues. These mutations, located in th e biotin carboxylase domain, provide a unique opportunity to analyze h ow natural occurring mutations affect PC function.