Crystallization and Crystallographic Analyses of Triosephosphate Isomerase from Naegleria gruberi

Resumo

The enzyme Triosephosphate Isomerase (TIM) from Naegleria gruberi is involved in the glycolytic pathway, in which it acts upon the conversion of dihydroxyacetone phosphate to D-glyceraldehyde 3-phosphate. Currently, the most similar TIM with 3D structure reported is from the plant Arabidopsis thaliana, which has 58% sequence identity with NgTIM. Recombinant NgTIM protein was expressed in E. coli BL21(DE3), purified by affinity and size exclusion chromatographies and crystallized by the vapor diffusion method. The best crystals were obtained in 3 different conditions, the one to provide the best dataset in ethanol 15% (V/V) and pentaerythritol propoxylate 40% (V/V). X ray diffraction data were collected for this crystal at 2.64 Å resolution and processed. It belongs to the space group P4122 and have unit cell parameters a = 79.70, c = 98.11 Å. The asymmetric unit contains one monomer, with VM of 2.68 Å3 Da-1 and a solvent content of 54.2 %.