SCREENING FOR POINT MUTATIONS IN EXON-10 OF THE LOW-DENSITY-LIPOPROTEIN RECEPTOR GENE BY ANALYSIS OF SINGLE-STRAND CONFORMATION POLYMORPHISMS - DETECTION OF A NONSENSE MUTATION FH(469-]STOP)

DNA from 40 unrelated familial hypercholesterolemia (FH) heterozygotes were subjected to analyses of single-strand conformation polymorphism s (SSCPs) of exon 10 of the low density lipoprotein receptor (LDLR) ge ne. Four different SSCP patterns were observed. The underlying mutatio ns were characterized by DNA sequencing. Three of the patterns represe nted the three genotypes of a recently described sense mutation in cod on 450. A method based upon the polymerase chain reaction (PCR) was de veloped to analyze this mutation. The frequencies of the wild-type (G at nucleotide 1413) and mutant (A at nucleotide 1413) alleles were 0.5 6 and 0.44, respectively. The fourth pattern was found in only one FH heterozygote and was caused by heterozygosity at nucleotide 1469 (G/A) . Nucleotide 1469 is the second base of codon 469Trp(TGG). The G-->A m utation changes this codon into the amber stop codon, and is referred to as FH469-->Stop. The mutant receptor consists of the amino terminal 468 amino acids. Because the truncated receptor has lost the membrane -spanning domain, it will not be anchored in the cell membrane. FH469- ->Stop destroys an AvaII restriction site, and this characteristic was used to develop a PCR method to establish its frequency in Norwegian FH subjects. Two out of 204 (1%) unrelated FH heterozygotes possessed the mutation.