Characterization of two Turkish beta-hexosaminidase mutations causing Tay-Sachs disease.
Two homoallelic mutations have recently been identified in the alpha-subunit of hexosaminidase A (EC 3.2.1.52) causing the infantile form of Tay-Sachs disease in Turkish patients. Both of these mutations, a 12 bp deletion (1096-1107 or 1098-1108 or 1099-1109) in exon 10 and a point mutation (G1362 to A, Gly454 to Asp) in exon 12, are located in the catalytic domain of the hexosaminidase alpha-chain. In order to determine whether these mutations affect the function of the catalytic domain or result in an instable protein, both mutant cDNAs were overexpressed in COS-1 cells. As judged by Western blotting, transfections of wild-type cDNA produced pro-alpha-chain and mature alpha-chain in parallel with a fivefold increase in cellular hexosaminidase activity using the synthetic substrate 4-methylumbelliferyl beta-N-acetylglucosamine 6-sulfate (MUGS). However, both mutants produced only pro-alpha-chains, although no mature form or detectable hexosaminidase activity towards two different synthetic substrates was observed. These data are consistent with the biochemical phenotype of infantile Tay-Sachs disease. We conclude that the overexpressed mutant pro-alpha-chains were misfolded and could not undergo further proteolytic processing to the active form of the enzyme in the lysosome.
Ozkara
Hatice Asuman
HA
Department of Biochemistry, Hacettepe University Faculty of Medicine, 06100 Ankara, Turkey. ozkara@ato.org.tu
Sandhoff
Konrad
K
D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000235:genetics; D054818:Hexosaminidase A; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010641:Phenotype; D017510:Protein Folding; D013661:Tay-Sachs Disease / Q000201:enzymology / Q000208:ethnology / Q000235:genetics*; D014162:Transfection; D014421:Turkey / Q000208:ethnology; D001619:beta-N-Acetylhexosaminidases / Q000737:chemistry / Q000235:genetics*
D018076:DNA, Complementary; D054818:Hexosaminidase A; D001619:beta-N-Acetylhexosaminidases