1.7 angstrom structure of the stabilized REIv mutant T39K. Application of local NCS restraints

The X-ray structure of the T39K mutant of the variable domain of a human im munoglobulin kappa light chain has been determined at room temperature to 1 .7 Angstrom resolution with a conventional R factor of 0.182. T39K crystall izes in the triclinic space group P1 [a = 35.4(1), b = 40.1(1), c = 43.1 (1 ) Angstrom, alpha = 66.9 (1), beta = 85.4 (1), gamma = 73.8 (1)degrees]. Th e unit-cell contains two monomers, related by a non-crystallographic twofol d arris, The use of a novel type of local non;crystallographic symmetry res traints on related isotropic displacement parameters and 1-4 distances as i ncorporated in the refinement program SHELXL improves the model and quality of the maps, but local differences between both monomers in areas subject to different packing contacts can still be observed. 12 overall anisotropic scaling parameters were refined, These may have compensated for the diffic ulties in accurately scaling single rotation axis image-plate data from a t riclinic crystal, because of the scarcity of common equivalent reflections. The final model has been used to perform a number of tests on anisotropic scaling, non-crystallographic symmetry, anisotropic refinement, determinati on of standard uncertainties and bulk solvent correction. It is remarkable that removal of the NCS restraints from the final model caused R-free to in crease. These tests clarify the strategies for optimum use of SHELXL for re finement at: medium as opposed to atomic resolution.