Tay-Sachs disease is an autosomal recessive genetic disorder resulting from mutation of the HEXA gene encoding the alpha-subunit of the lysosomal enzyme, beta-N-acetylhexosaminidase A (ref. 1). A relatively high frequency of carriers (1/27) of a lethal, infantile form of the disease is found in the Ashkenazi Jewish population, but it is not yet evident whether this has resulted from a founder effect and random genetic drift or from a selective advantage of heterozygotes. We have identified a single-base mutation in a cloned fragment of the HEXA gene from an Ashkenazi Jewish patient. This change, the substitution of a C for G in the first nucleotide of intron 12 is expected to result in defective splicing of the messenger RNA. A test for the mutant allele based on amplification of DNA by the 'polymerase chain rection and cleavage of a DdeI restriction site generated by the mutation revealed that this case and two other cases of the Ashkenazi, infantile form of Tay-Sachs disease are heterozygous for two different mutations. The occurrence of multiple mutant alleles warrants further examination of the selective advantage hypothesis.
We have reported previously the isolation and partial characterization of a 215-kilodalton (Kd) phosphomannosyl binding protein from bovine liver membranes [3,9]. In the present studies evidence is presented that the binding protein is an aggregate. Four N-terminal amino acids were detected, and the complex could be dissociated into subunits. Bovine liver membranes were extracted with the detergent, Zwittergent, in the presence of protease inhibitors. The extract was subjected to affinity chromatography on phosphomannan-Sepharose 4B, and proteins with apparent Mr values of 215 and 57 Kd were eluted with mannose 6-phosphate. As reported previously, extraction with Triton X-100 yielded only the higher molecular weight material. When the binding protein was incubated at 4 degrees C in the presence of Zwittergent TM 3-14 the 215-Kd form slowly dissociated into smaller subunits; after two months, the major species had an apparent Mr of 57 Kd. The subunits derived from the binding protein were recognized by antiserum raised against purified binding protein. Dissociation of the binding protein by Zwittergent was enhanced by incubation at 37 degrees C, the presence of dithiothreitol, and low pH values. The subunit mixture enriched in the 57-Kd subunit had a lowered ability to bind ligands containing the phosphomannosyl recognition marker. Binding was partially restored (greater than 48% of the initial value) when dissociated receptor was back exchanged with Triton X-100.
Phosphomannosyl receptor (PMR) isolated from Syrian hamster liver was purified to apparent homogeneity by affinity chromatography and one-dimensional PAGE.
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