Dynamic light-scattering and preliminary crystallographic studies of the sensor domain of the haem-based oxygen sensor FixL from Rhizobium meliloti

FixL is a transmitter protein in a two-component system which acts as an oxygen sensor when the symbiotic Rhizobia resides in root nodules of host plants. The oxygen-sensor domain of Rhizobium meliloti FixL (RmFixLH) was purified by His-tag affinity and isoelectronic focusing chromatographies, without the use of gel-filtration chromatography. Dynamic light-scattering measurements of RmFixLH thus obtained revealed it to be monodispersive and present as a homodimer in solution. A single crystal of RmFixLH in the met (Fe³⁺) form was grown in 100 mM acetic acid/NaOH buffer at pH 4.6 in the presence of 200 mM ammonium acetate, using 40%(w/­v) PEG 4000 as a precipitant. A crystal of the ferrous CO form of RmFixLH was also prepared by reduction of the met crystal with Na₂S₂O₄ in an atmosphere of CO. The crystals (0.2 × 0.05 × 0.01 mm) belong to the monoclinic system (C2) with unit-cell parameters a = 60.94, b = 37.44, c = 54.14 Å, β = 115.29° and diffract X-rays to 1.7 Å resolution at station BL44B2 of SPring-8, Japan. Bijvoet difference Patterson maps show a clear peak corresponding to the haem iron in RmFixLH.