Cloning and expression of the defective genes in delta-aminolevulinate dehydratase porphyria: compound heterozygosity in this hereditary liver disease.

Cloning and expression of the defective genes for ALAD from a patient with inherited ADP were carried out. Two separate point mutations, termed G1 and G2, resulting in a single amino acid change in each ALAD allele, were identified. The G1 mutation (C718-->T) occurred in the allele within the substrate-binding site, producing an Arg240-->Trp substitution; the G2 mutation (G820-->A) occurred downstream of this site in the other allele, resulting in an Ala274-->Thr substitution. Using RT-PCR, the mother, the brother, and the sister were shown to have the G1 defect. Expression of the G1 cDNA in CHO cells produced ALAD protein with little activity; the G2 cDNA produced the enzyme with approximately 50% normal activity. Pulse-labeling studies demonstrated that the G1 enzyme had a normal half-life, while the G2 enzyme had a markedly decreased half-life. These data thus define two separate point mutations, one in each ALAD allele, as well as the altered properties of the two enzymic proteins encoded by the mutant genes in this patient.