A novel mutation of delta-aminolaevulinate dehydratase in a healthy child with 12% erythrocyte enzyme activity

Cloning, expression and phenotype studies of the defective gene for delta-a minolaevulinate dehydratase (ALAD) in a family with an asymptomatic girl wh o had ALAD deficiency were carried out. The proband was identified by neona tal ALAD screening, and had erythrocyte ALAD activity at 12% of the normal control. She was heterozygous for ALAD deficiency, which was inherited from her father. Nucleotide sequence analysis of the cloned ALAD cDNA revealed C-36 to G and T-168 to C mutations on the same allele. The former mutation resulted in F12L substitution, whereas the latter was a silent mutation. Al l family members who had decreased ALAD activity had the same mutation. Exp ression of the mutant ALAD cDNA in Chinese hamster ovary cells produced an ALAD protein without significant enzyme activity. Additionally, the mutant ALAD cDNA which encodes F12L substitution produced an aberrant migration pa ttern in polyacrylamide gel electrophoresis under denaturing conditions. Th is finding probably reflects an abnormal folding of the F12L protein, since the mutation occurred in the al helix of the N-terminal arm of the enzyme, which is involved in the extensive quaternary interactions among the subun its, This is also the first report of ALAD gene mutation in an asymptomatic subject.