Reexamination of alcohol dehydrogenase structural mutants in Drosophila using protein blotting.
Using protein blotting and an immuno-overlay procedure, we have reexamined the cross-reacting material produced by ADH null-activity mutants generated with ethyl methanesulfonate (EMS). Of the 13 mutants, 11 have an immunodetectable polypeptide of wild-type size. The native and urea denatured isoelectric points (pI) establish that 7 of 13 of the mutations have no effect on protein charge. The electrophoretic mobilities of each variant on increasing percent acrylamide gels (Ferguson analysis), reveal that 9 of the 11 immunodetectable mutants have retained the ability to form dimers under native conditions. None of the inactive mutant proteins has the ability to form the "adduct-bound" isozyme. We have found no correlation between protein pI and in vivo stability. The observed frequencies of specific charge class alterations do not dispute the propensity of G:A transitions previously found for EMS mutagenesis.
D000426:Alcohol Dehydrogenase / Q000235:genetics* / Q000302:isolation & purification; D000818:Animals; D004331:Drosophila melanogaster / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005020:Ethyl Methanesulfonate / Q000494:pharmacology*; D005796:Genes / Q000187:drug effects*; D005838:Genotype; D009154:Mutation
D005020:Ethyl Methanesulfonate; D000426:Alcohol Dehydrogenase