Purification crystallization and preliminary X-ray diffraction studies of the three components of the toluene 23-dioxygenase enzyme system

摘要

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 (ReductaseTOL), a ferredoxin (FerredoxinTOL) and a Rieske non-heme iron dioxygenase (OxygenaseTOL). The three components and the apoenzyme of the dioxygenase (apo-OxygenaseTOL) were overexpressed, purified and crystallized. ReductaseTOL diffracts to 1.8 Å and belongs to space group P41212, with unit-cell parameters a = b = 77.1, c = 156.3 Å. FerredoxinTOL 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-OxygenaseTOL and OxygenaseTOL 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.