STRUCTURE OF COPPER-SUBSTITUTED AND OXALATE-SUBSTITUTED HUMAN LACTOFERRIN AT 2.0 ANGSTROM RESOLUTION

The three-dimensional structure of human dicupric monooxalate lactoferrin, Cu2oxLf, has been determined to 2.0 angstrom resolution, using X-ray diffraction data collected by diffractometry to 2.5 angstrom resolution, and oscillation photography on a synchrotron source to 2.0 angstrom resolution. Difference electron-density maps calculated between CU2oxLf and both dicupric lactoferrin, Cu2Lf, and diferric lactoferrin, Fe2Lf, showed that the oxalate had replaced a carbonate in the C-terminal binding site, and that, relative to Cu2Lf, there were no significant differences in the N-terminal site. The structure was then refined crystallographically by restrained least-squares methods. The final model, in which the r.m.s. deviation in bond distances is 0.017 angstrom, contains 5314 protein atoms (691 residues), two Cu2+ ions, one bicarbonate ion, one oxalate ion, 325 solvent molecules and one sugar residue. The crystallographic R factor of 0.193 is for 46 134 reflections in the range 8.0 to 2.0 angstrom resolution. The oxalate ion is coordinated to copper in a 1,2-bidentate fashion, and the added bulk of the anion results in the rearrangement of the side chains of nearby arginine and tyrosine residues. No other major alterations in the molecule can be observed, the overall protein structure being the same as that for Cu2Lf and Fe2Lf.