CRYSTAL-STRUCTURE OF USTILAGO-SPHAEROGENA RIBONUCLEASE U-2 AT 1.8-ANGSTROM RESOLUTION

The crystal structure of purine-specific ribonuclease (RNase) U-2 from Ustilago sphaerogena has been solved by the molecular replacement method using RNase T-1 as a search model. The structure, with 114 amino acid residues, 141 water molecules, and a sulfate ion, is refined to an R factor of 0.143 at 1.8 Angstrom resolution. As evidenced by the electron densities, residues 49 and 50 are revised to Glu 49 and Asp 50, respectively, and also Asp 45 is identified as a beta-isomerized form to L-isoaspartate with a beta-peptide Linkage. RNase U-2 consists of a beta-hairpin at residues from 7 to 14, a 4.4-turn alpha-helix from 16 to 32, a central beta-sheet with five strands, and a protruding beta-turn from 74 to 77. As for the catalytic site residues, His 41, Glu 62, and Arg 85 are located as constituents of the central beta-sheet, and Tyr 39 and His 101 are situated at either end of the beta-sheet. The side chains of Tyr 39, Glu 62, Arg 85, and His 101 are hydrogen-bonded to the sulfate ion which marks the RNA phosphate position. Though the side chain of His 41 is pointing away from the sulfate, small conformational adjustments of His 41 enable the side chain to interact with either the phosphate or the ribose group of RNA. The loop region from Tyr 44 to Asp 50 is ascribed to the base recognition site where Glu 49 is involved in adenine recognition. beta-Isomerized Asp 45 suggests that this region is conformationally flexible and alterable.