The crystal structure of subtilisin Carlsberg in anhydrous dioxane and its comparison with those in water and acetonitrile.

The x-ray crystal structure of the serine protease subtilisin Carlsberg in anhydrous dioxane has been determined to 2.6-A resolution. The enzyme structure is found to be nearly indistinguishable from the structures previously determined in water and acetonitrile. Small changes in the side-chain conformations between the dioxane and water structures are of the same magnitude as those observed between two structures in different aqueous systems. Seven enzyme-bound dioxane molecules have been detected, each potentially forming at least one hydrogen bond with a subtilisin hydrogen-bond donor or bound water. Two of the bound dioxane molecules are in the active-site region, one in the P2 and another bridging the P1' and P3' pockets. The other five dioxane molecules are located on the surface of subtilisin at interprotein crystal contacts. The locations of the bound solvent in the dioxane structure are distinct from those in the structures in acetonitrile and in water.