Deficiency of propionyl-CoA carboxylase (PCC) results in propionic acidemia, an 
autosomal recessive disorder characterized by ketoacidosis sufficiently severe 
to cause neonatal death. PCC is involved in the catabolism of branched-chain 
amino acids, odd-chain fatty acids, and cholesterol. The enzyme is a 
biotin-dependent mitochondrial protein composed of two heterologous subunits 
arranged into an 800-kDa alpha(6 )beta(6) dodecameric structure. Approximately 
60 mutations have been reported in the nuclear genes PCCA and PCCB that encode 
the two PCC subunits. The vast majority of these mutations have not been 
examined at the protein level. We present an initial characterization of 13 
mutations located in exons 1, 3-7, and 12-14 of PCCB. After expression in E. 
coli, these recombinant mutant enzymes were analyzed for stability, 
biotinylation, alpha-beta subunit interaction, and activity. Our results show a 
functional dichotomy in these PCCB mutations with some mutants (R44P, S106R, 
G131R, G198D, V205D, I408del, and M442T) capable of varying degrees of assembly 
but forming catalytically inactive PCC proteins. Other PCCB mutants (R165W, 
E168K, D178H, P228L, and R410W) that are PCC deficient in patient-derived 
fibroblasts, were found to be capable of expressing wild-type level PCC activity 
when assembled in our chaperone-assisted E. coli expression system. This result 
indicates that these mutations exert their pathogenic effect due to an inability 
to assemble correctly in patients' cells. This initial screen has identified a 
range of mutant PCC proteins that are sufficiently stable to be purified and 
subsequently used for structure-function analysis to further elucidate the 
complex relationship between genotype and phenotype in propionic acidemia.