Characterization of two Turkish β-hexosaminidase mutations causing Tay-Sachs disease

Two homoallelic mutations have recently been identified in the α-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 α-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-α-chain and mature α-chain in parallel with a fivefold increase in cellular hexosaminidase activity using the synthetic substrate 4-methylumbelliferyl β-N-acetylglucosamine 6-sulfate (MUGS). However, both mutants produced only pro-α-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-α-chains were misfolded and could not undergo further proteolytic processing to the active form of the enzyme in the lysosome.