Expression, purification, crystallization and preliminary crystallographic analysis of recombinant pteridine reductase of Trypanosoma cruzi

Abstract

The recombinant version of Trypanosoma cruzi pteridine reductase was expressed in Escherichia coli, purified to homogeneity from the soluble fraction of bacterial extract by metal-chelate affinity chromatography and crystallized in the presence of the cofactor (NADPH) and an inhibitor (methotrexate) at 295 K using sodium acetate as precipitant. The crystals are trigonal, belonging to space group P3(1) (or P3(2)), with unit-cell parameters a = 74.35, c = 179.96 A under cryogenic conditions. The asymmetric unit contains a tetramer, with a corresponding V(M) of 2.3 A(3) Da(-1)and a solvent content of 46%. Native data have been collected to 2.1 A resolution using Cu Kalpha X-rays.