Molecular analysis of patients with holocarboxylase synthetase deficiency by fluorescence-based single-strand conformation polymorphism (SSCP) analysis

Holocarboxylase synthetase (HCS) deficiency is an autosomal recessive disorder characterized by combined organic aciduria, metabolic ketoacidosis, and dermatitis. These clinical symptoms are dramatically improved by administration of biotin. We have previously identified two mutations (L237P and delG1067) that were frequent in Japanese patients. In order to detect mutations of three German patients and one French patient, we used fluorescence-based PCR-SSCP analysis on 12 overlapping HCS cDNA fragments. Two abnormal migration patterns were detected in fragments 5, 6, and 8, and one abnormal pattern was detected in fragments 9 and 12. Sequencing analysis of cDNA revealed five new mutations; a C to T substitution at nt. position 1121 (Ser-Ser) in patients 1 and 3, a T to A substitution at nt. position 1285 (Val->Glu) in patient 1, a single T deletion at nt. position 1876 resulting in a stop codon in patient 4, 68-base deletion between nt. positions 1740 and 1807 also in patient 4, and a single C deletion at nt. position 2279 followed by a stop codon in patient 2. No mutations were detected in 2 of 8 aberrant patterns. To identify other mutations, SSCP analysis on gels containing 5% glycerol is now in progress.