Phenylalanine hydroxylase. Evidence that the enzyme from human liver might not be a phosphoprotein.

Phenylalanine hydroxylase from fresh human liver was purified to homogeneity with a 60% yield by a three steps procedure involving hydrophobic chromatography on Phenyl-Sepharose, ion exchange chromatography on DEAE-Cellulose and High Performance gel permeation chromatography. The purified native enzyme had an estimated molecular weight of 165,000. It gave a single band on Sodium Dodecylsulfate polyacrylamide gel electrophoresis under an estimated molecular weight of 55,000. Neither the purified human enzyme, nor that present in a liver extract could be activated under phosphorylating conditions. Moreover, the purified human liver phenylalanine hydroxylase was found to be devoid of protein-bound phosphate and no phosphate could be incorporated from [32P]-ATP in the presence of cyclic-AMP - dependent protein kinase. These data suggest that phenylalanine hydroxylase from human liver, unlike that of rat liver, might not be a phosphoprotein.