Application of a copper blotting method to the study of Menkes disease |
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Authors: | Stephen G Kaler William A Gahl |
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Institution: | (1) Section on Human Biochemical Genetics, Human Genetics Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD |
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Abstract: | Menkes disease is an X-linked recessive disorder of copper metabolism. Deficient quantity or functional activity of a molecule
involved in intracellular copper transport is believed to represent the basic defect. We applied an in vitro copper binding
assay (copper blotting) to tissue proteins from Menkes patients and controls to evaluate differences in copper-binding. Proteins
were separated by SDS-PAGE, electrotransferred to nitrocellulose, and probed with67CuCl2. Copper-binding polypeptides were visualized by autoradiography. No major differences were observed between a Menkes patient
and control subjects in copper blots of post-mortem liver, kidney, or brain—tissues affected clinically by the disturbance
of copper metabolism in Menkes disease. We also applied the copper blotting technique to fibroblast proteins from an affected
female in whom the gene responsible for Menkes disease is interrupted by a chromosomal translocation, and detected no differences
in copper-binding proteins relative to normal controls. These experiments suggest that the gene product defective in Menkes
disease is not detectable in copper blots, either because normal tissue levels are below the limits of detection of this method,
or because the molecule involved does not bind copper under these conditions. |
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Keywords: | Menkes disease 67CuCl2 copper blotting |
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