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A glycine 250 → aspartate substitution in the α‐subunit of hexosaminidase a causes juvenile‐onset Tay‐Sachs disease in a Lebanese‐Canadian family
Author(s) -
Trop Isabelle,
Kaplan Feige,
Brown Charlotte,
Mahuran Don,
Hechtman Peter
Publication year - 1992
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.1380010106
Subject(s) - biology , complementary dna , microbiology and biotechnology , exon , mutant , mutation , transfection , gene , genetics
The mutation causing juvenile Tay‐Sachs disease (TSD) in two sibs of Lebanese‐Maronite origin is described. An mRNA‐containing extract of cultured fibroblasts obtained from one of the probands was used as a template to amplify the coding sequence of the hexoaminidase A (Hex A) α‐subunit. Sequencing of amplified cDNA fragments revealed a single alteration, guanine to adenine at nt 749 creating a G250D mutation. The mutation introduces a new recognition site for the restriction enzyme Eco RV, permitting identification of heterozygotes for this allele following PCR amplification and Eco RV digestion of exon 7 sequences from genomic DNA templates. In order to test the effect of this substitution, an in vitro mutagenized cDNA construct was introduced into a mammalian expression vector and transfected into monkey Cos‐1 cells separately or along with a β‐cDNA expression vector. When the mutant α‐cDNA was the only gene introduced into COS cells no enzymatic activity above endogenous COS cell activity was detected. Cotransfection of normal α‐cDNA and β‐cDNA followed by immunoprecipitation of human Hex A resulted in 20‐fold increase in the ratio between positive and negative (mock transfection) control values. This allowed the detection of some residual activity (12% of the positive control) when the mutant α‐cDNA replaced its wild‐type counterpart. The predicted protein environment in which the mutation occurs is compared to that of the adult‐onset Tay‐Sachs disease mutation caused by a Gly 269 → Ser substitution in exon 7. We suggest that the more severe clinical phenotype linked to the Gly 250 → Asp mutation may be explained by the less conservative substitution of the large, charged aspartate residue as compared to the small neutral serine, and the placement of the Gly 250 in a hydrophobic section of an α‐helix. © 1992 Wiley‐Liss, Inc.