DOMAIN-STRUCTURE AND STABILITY OF HUMAN PHENYLALANINE-HYDROXYLASE INFERRED FROM INFRARED-SPECTROSCOPY

We have studied the conformation and thermal stability of recombinant human phenylalanine hydroxylase (hPAH) and selected truncated forms, c orresponding to distinct functional domains, by infrared spectroscopy. The secondary structure of wild-type hPAH mas estimated to be 48% alp ha-helix, 28% extended structures, 12% beta-turns and 12% non-structur ed conformations. The catalytic C-terminal domain (residues 112-452) h olds most of the regular secondary structure elements, whereas the reg ulatory N-terminal domain (residues 2-110) adopts mainly an extended a nd disordered, flexible conformation. Thermal stability studies of the enzyme forms indicate the existence of interactions between the two d omains. Our results also demonstrate that the conformational events in volved in the activation of hPAH by its substrate (L-Phe) are mainly r elated to changes in the tertiary/quaternary structure. The activating effect of phosphorylation, however, affects the secondary structure o f the N-terminal domain of the protein. (C) 1998 Federation of Europea n Biochemical Societies.