Theoretical study of structure of catalytic copper site in nitrite reductase

The catalytic copper site in nitrite reductase contains a Cu2+ ion bound to three histidine (His) ligands and a solvent molecule. Sites from various sources show a conspicuous variation in the structure. In some proteins, it is close to tetrahedral (even more so than are blue copper proteins), whereas in other proteins, it has a structure more similar to that expected for a type 2 copper site. We have studied this site with a number of theoretical methods, ranging from vacuum optimizations, combined quantum and molecular mechanics (QM/MM) optimization, quantum refinement (X-ray crystallography supplemented by quantum chemical calculations), and accurate energy calculations. We show that the difference in the structure arises from a movement of the solvent molecule and that this movement is determined by a compromise between its hydrogen bond interactions and the intrinsic preferences of the copper site. If the solvent molecule is deprotonated, the two structures have a similar energy, whereas if it is protonated, the more tetrahedral structure is energetically favorable. Neither of the structures involves a pi interaction as in the blue copper proteins; instead, both are strongly distorted tetragonal structures with sigma bonds to all four ligands. We have also examined the position of hydrogen atoms shared between second-sphere carboxylate groups and the first-sphere solvent molecule and one of the His ligands. In the oxidized state, the structure with the solvent deprotonate(d) but the His residue protonated seems to be most stable. (c) 2004 Wiley Periodicals, Inc.