Structure of mutant human carbonmonoxy­hemoglobin C (βE6K) at 2.0 Å resolution

Previous studies have demonstrated that in vitro crystallization of R-state liganded hemoglobin C (HbC), a naturally occurring mutant human hemoglobin (βE6K), in high-phosphate buffer solutions provides a potential model system for the intracellular crystallization of HbC associated with chronic hemolytic anemia in CC disease. The first high-resolution crystal structure of liganded HbC is reported here. HbC was crystallized from high phosphate and the structure of the carbonmonoxy-liganded R-state form was refined at 2.0 Å resolution. Crystals exhibit diffraction consistent with the tetragonal space group P4₁2₁2, with unit-cell parameters a  = 54.16, c = 195.30 Å. The structure was solved by difference Fourier techniques and refinement by simulated annealing and restrained least-squares yielded a final R of 0.183 and an R_free of  0.238 for all 19 382 unique reflections. The side chain of βK6 exhibits very weak electron density consistent with significant mobility within the crystalline lattice. The highly dynamic nature of the side chain could potentially support a number of specific polar interactions that might reduce the barrier to crystallization and thus result in enhanced crystallization kinetics for HbC relative to HbA. Specifically, the NZ atom of the BK6 side chain could participate in an amino–aromatic hydrogen bond with the π-electron cloud of βH116 in a symmetry-related tetramer. βK6 NZ might also interact with the main-chain carbonyl O atom of βH117 and the carboxylate group of βE22 from a symmetry-related tetramer.