Characterization of propionic acidemia mutations A113T and I139T using E. coli biotin carboxylase

Abstract

Propionic Acidemia (PA) is an inborn error of metabolism caused by a deficiency of propionyl‐CoA carboxylase (PCC). PCC is a biotin dependent carboxylase involved in odd chain fatty acid oxidation and branched chain amino acid catabolism. Two mutations in the αsubunit of PCC that result in PA are A113T and I139T. The homodimeric biotin carboxylase (BC) subunit of E. coli acetyl‐CoA carboxylase is a model system for studying these mutations. Sequence alignments revealed the mutations A113T and I139T in PCC are homologous to A77T and I103T in BC. These residues are very close in proximity lying in adjacent β‐sheets far from the active site. Kinetic analysis was carried out for WT and both mutants holding biotin and bicarbonate constant while varying ATP. The results showed the I103T had no change in Km for ATP and only a 3‐fold lower Vmax with respect to WT. In contrast, the A77T mutant exhibited negative cooperativity with respect to ATP. This result provides further evidence for communication between the two subunits of E. coli BC and insight into the molecular pathogenesis of this mutation.