The molecular basis of 3-methylcrotonylglycinuria, a disorder of leucine catabolism

3-Methylcrotonylglycinuria is an inborn error of leucine catabolism and has a recessive pattern of inheritance that results from the deficiency of 3-m ethylcrotonyl-CoA carboxylase (MCC). The introduction of tandem mass spectr ometry in newborn screening has revealed an unexpectedly high incidence of this disorder, which, in certain areas, appears to be the most frequent org anic aciduria. MCC, an heteromeric enzyme consisting of alpha (biotin-conta ining) and beta subunits, is the only one of the four biotin-dependent carb oxylases known in humans that has genes that have not yet been characterize d, precluding molecular studies of this disease. Here we report the charact erization, at the genomic level and at the cDNA level, of both the MCCA gen e and the MCCB gene, encoding the MCC alpha and MCC beta subunits, respecti vely. The 19-exon MCCA gene maps to 3q25-27 and encodes a 725-residue prote in with a biotin attachment site; the 17-exon MCCB gene maps to 5q12-q13 an d encodes a 563-residue polypeptide. We show that disease-causing mutations can be classified into two complementation groups, denoted "CGA" and "CGB. " We detected two MCCA missense mutations in CGA patients, one of which lea ds to absence of biotinylated MCC alpha. Two MCCB missense mutations and on e splicing defect mutation leading to early MCC beta truncation were found in CGB patients. A fourth MCCB mutation also leading to early MCC beta trun cation was found in two nonclassified patients. A fungal model carrying an mccA null allele has been constructed and was used to demonstrate, in vivo, the involvement of MCC in leucine catabolism. These results establish that 3-methylcrotonylglycinuria results from loss-of-function mutations in the genes encoding the alpha and beta subunits of MCC and complete the genetic characterization of the four human biotin-dependent carboxylases.