title: W474C amino acid substitution affects early processing of the alpha-subunit of beta-hexosaminidase A and is associated with subacute G(M2) gangliosidosis.
abstract: Mutations in the HEXA gene, encoding the alpha-subunit of beta-hexosaminidase A (Hex A), that abolish Hex A enzyme activity cause Tay-Sachs disease (TSD), the fatal infantile form of G(M2) gangliosidosis, Type 1. Less severe, subacute (juvenile-onset) and chronic (adult-onset) variants are characterized by a broad spectrum of clinical manifestations and are associated with residual levels of Hex A enzyme activity. We identified a 1422 G-->C (amino acid W474C) substitution in the first position of exon 13 of HEXA of a non-Jewish proband who manifested a subacute variant of G(M2) gangliosidosis. On the second maternally inherited allele, we identified the common infantile disease-causing 4-bp insertion, +TATC 1278, in exon 11. Pulse-chase analysis using proband fibroblasts revealed that the W474C-containing alpha-subunit precursor was normally synthesized, but not phosphorylated or secreted, and the mature lysosomal alpha-subunit was not detected. When the W474C-containing alpha-subunit was transiently co-expressed with the beta-subunit to produce Hex A (alphabeta) in COS-7 cells, the mature alpha-subunit was present, but its level was much lower than that from normal alpha-subunit transfections, although higher than in those cells transfected with an alpha-subunit associated with infantile TSD. Furthermore, the precursor level of the W474C alpha-subunit was found to accumulate in comparison to the normal alpha-subunit precursor levels. We conclude that the 1422 G-->C mutation is the cause of Hex A enzyme deficiency in the proband. The resulting W474C substitution clearly interferes with alpha-subunit processing, but because the base substitution falls at the first position of exon 13, aberrant splicing may also contribute to Hex A deficiency in this proband.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Manitoba, Winnipeg, Canada.
authors.forename: E
authors.initials: E
authors.lastname: Petroulakis
authors.forename: Z
authors.initials: Z
authors.lastname: Cao
authors.forename: J T
authors.initials: JT
authors.lastname: Clarke
authors.forename: D J
authors.initials: DJ
authors.lastname: Mahuran
authors.forename: G
authors.initials: G
authors.lastname: Lee
authors.forename: B
authors.initials: B
authors.lastname: Triggs-Raine
mesh_terms: D000293:Adolescent; D017668:Age of Onset; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D016521:Electrophoresis, Gel, Pulsed-Field; D005091:Exons; D005347:Fibroblasts / Q000201:enzymology; D054818:Hexosaminidase A; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017386:Sequence Homology, Amino Acid; D013661:Tay-Sachs Disease / Q000201:enzymology / Q000235:genetics*; D014162:Transfection; D001619:beta-N-Acetylhexosaminidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D054818:Hexosaminidase A; D001619:beta-N-Acetylhexosaminidases