Phenylketonuria (PKU) is a metabolic disorder due primarily to mutations in the PAH gene that impair both phenylalanine hydroxylase activity and disposal of l-phenylalanine from the normal diet. Excess phenylalanine is toxic to cognitive development and a low-phenylalanine diet prevents mental retardation, but it is a difficult therapeutic option. Previous studies with recombinant phenylalanine ammonia-lyase, PAL, demonstrated pharmacologic and physiologic proofs of principle for PAL as an alternative therapy for PKU but its immunogenicity was problematic. From a series of formulations of linear and branched polyethylene glycols chemically conjugated to PAL, we have created a parenteral therapeutic agent for PKU treatment. All the pegylated molecules were fully characterized in vitro and the most promising formulations were then tested in vivo in the PKU mouse model. The linear 20-kDa PEG–PAL combination abolished in vivo immunogenicity after repeated challenge while retaining full catabolic activity against phenylalanine, suggesting potential as a novel PKU therapeutic.