title: Structural comparison of bacterial and human iron-dependent phenylalanine hydroxylases: similar fold, different stability and reaction rates.
abstract: Structure determination of bacterial homologues of human disease-related proteins provides an efficient path to understanding the three-dimensional fold of proteins that are associated with human diseases. However, the precise locations of active-site residues are often quite different between bacterial and human versions of an enzyme, creating significant differences in the biological understanding of enzyme homologs. To study this hypothesis, phenylalanine hydroxylase from a bacterial source has been structurally characterized at high resolution and comparison is made to the human analog. The enzyme phenylalanine hydroxylase (PheOH) catalyzes the hydroxylation of l-phenylalanine into l-tyrosine utilizing the cofactors (6R)-l-erythro-5,6,7,8 tetrahydrobiopterin (BH(4)) and molecular oxygen. Previously determined X-ray structures of human and rat PheOH, with a sequence identity of more than 93%, show that these two structures are practically identical. It is thus of interest to compare the structure of the divergent Chromobacterium violaceum phenylalanine hydroxylase (CvPheOH) ( approximately 24% sequence identity overall) to the related human and rat PheOH structures. We have determined crystal structures of CvPheOH to high resolution in the apo-form (no Fe-added), Fe(III)-bound form, and 7,8-dihydro-l-biopterin (7,8-BH(2)) plus Fe(III)-bound form. The bacterial enzyme displays higher activity and thermal melting temperature, and structurally, differences are observed in the N and C termini, and in a loop close to the active-site iron atom.
authors.affiliation: The Scripps Research Institute, Department of Molecular Biology, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
authors.forename: Heidi
authors.initials: H
authors.lastname: Erlandsen
authors.forename: Joo Y
authors.initials: JY
authors.lastname: Kim
authors.forename: Marianne G
authors.initials: MG
authors.lastname: Patch
authors.forename: Andrew
authors.initials: A
authors.lastname: Han
authors.forename: Alon
authors.initials: A
authors.lastname: Volner
authors.forename: Mahdi M
authors.initials: MM
authors.lastname: Abu-Omar
authors.forename: Raymond C
authors.initials: RC
authors.lastname: Stevens
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism; D020134:Catalytic Domain / Q000235:genetics; D002861:Chromobacterium / Q000201:enzymology* / Q000235:genetics; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D006801:Humans; D007501:Iron / Q000378:metabolism; D007700:Kinetics; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010651:Phenylalanine Hydroxylase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010661:Phenylketonurias / Q000201:enzymology / Q000235:genetics; D011487:Protein Conformation; D017510:Protein Folding; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
chemical_list: D008670:Metals; D011994:Recombinant Proteins; D001708:Biopterins; D007501:Iron; D010651:Phenylalanine Hydroxylase; C003402:sapropterin