We have examined 11 previously described cultured rat
hepatoma mutants with absent or reduced
phenylalanine hydroxylase activity (Choo and Cotton, 1977). Immunological and electrophoretic methods failed to detect any structurally altered
protein in these mutants. In nine independently isolated revertants from four different mutants, wild-type
protein was regained (or accentuated). This evidence suggests that the mutation involved in these mutants is most likely to be regulatory in nature. These studies have provided three reasons for believing that in cultured rat
hepatoma cells one gene codes for a single
polypeptide chain, a number of which combine to form the active
phenylalanine hydroxylase multimer: (1) Analysis of the purified
protein by two-dimensional electrophoresis revealed only a single
polypeptide chain. (2) This
polypeptide was diminished or undetectable in
crude extracts of 11 independently isolated mutants with absent of reduced activity. (3) In none of these 11 mutants was the
polypeptide we have designated to be
phenylalanine hydroxylase present at normal levels, as would be expected if the mutation were at another locus responsible for a possible second subunit.