Crystallization and X-ray analysis of a bacterial non-haem iron-containing phenylalanine hydroxylase from the Gram-negative opportunistic pathogen Pseudomonas aeruginosa.
Monooxygenases are frequently involved in the pathways that mediate the pivotal role of microorganisms in recycling carbon from the environment. A structural study of a monooxygenase from Pseudomonas aeruginosa that was identified as a phenylalanine hydroxylase has been initiated. The single-domain monomeric protein harbours a non-haem iron at the active site. The sequence identity to the catalytic domains of tyrosine and tryptophan hydroxylases suggests that the enzyme is not restricted to the substrate phenylalanine alone. Here, the cloning, purification and crystallization of native and SeMet-labelled P. aeruginosa phenylalanine hydroxylase are reported. Crystals grew in space group P6(1), with unit-cell parameters a = b = 210.5, c = 100.7 A, and diffracted to a d spacing of 2.0 A. Crystals of SeMet-labelled protein were used to collect a three-wavelength multiple anomalous dispersion (MAD) data set around the Se K edge.
Ekström
Fredrik
F
Umeå Centre for Molecular Pathogenesis, UCMP, Umeå University, SE-901 87, Umeå, Sweden.
Stier
Gunter
G
Eaton
Julian T
JT
Sauer
Uwe H
UH
D001426:Bacterial Proteins / Q000737:chemistry; D003001:Cloning, Molecular; D003460:Crystallization / Q000379:methods; D007501:Iron; D010651:Phenylalanine Hydroxylase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D012645:Selenomethionine; D014961:X-Ray Diffraction / Q000379:methods
D001426:Bacterial Proteins; D012645:Selenomethionine; D007501:Iron; D010651:Phenylalanine Hydroxylase