Purification, crystallization and preliminary X-ray diffraction studies of the three components of the toluene 2,3-dioxygenase enzyme system

Pseudomonas putida F1 can grow with toluene as its sole source of carbon and energy. The initial reaction of the degradation of toluene is catalyzed by a three-component toluene dioxygenase enzyme system consisting of a reductase (Reductase_TOL), a ferredoxin (Ferredoxin_TOL) and a Rieske non-heme iron dioxygenase (Oxygenase_TOL). The three components and the apoenzyme of the dioxygenase (apo-Oxygenase_TOL) were overexpressed, purified and crystallized. Reductase_TOL diffracts to 1.8 Å and belongs to space group P4₁2₁2, with unit-cell parameters a = b = 77.1, c = 156.3 Å. Ferredoxin_TOL diffracts to 1.2 Å and belongs to space group P2₁, with unit-cell parameters a = 30.5, b = 52.0, c = 30.95 Å, β = 113.7°. Apo-Oxygenase_TOL and Oxygenase_TOL diffract to 3.2 Å and belong to space group P4₃32, with unit-cell parameters a = 235.9 Å and a = 234.5 Å, respectively.