Refinement of the crystal structure of wheat germ agglutinin isolectin 2 at 1.8Åresolution

Abstract

The crystal structure of wheat germ agglutinin isolectin 2 has been refined by the restrained least-squares method of Hendrickson & Konnert (1980). The asymmetric unit of the C2 crystals contains two chemically identical protomers related by a non-crystallographic 2-fold screw operation. A total of 2290 protein atoms and 186 ordered water sites refined to a final R-factor of 0.179 and an average B-value of 21.6Å 2, using 54% (15,601) of the total possible number of reflections in the resolution range 8 to 1.8Åwith F o > 3σ(F o). The final model conforms to stereochemically correct bond distances and angles with root-mean-square (r.m.s.) values of 0.018Åand 3.3 °, respectively. Accuracy of this model is estimated to be 0.20Åon the basis of a Luzzati plot. Main-chain atomic positions in the two independent promoters, designated I and II, agree with an r.m.s. deviation of 0.30Å(0.58Åfor all atoms), indicating identical backbone conformation. The largest discrepancies are seen at flexible surface residues. One error was detected in the amino acid sequence at position 41 (Ser), which refined satisfactorily as a Trp. Loss of electron density for residue A171 during the course of refinement suggests either disorder or absence of this C-terminal residue. residue domains (A, B, C and D), was analyzed in terms of dihedral angles, backbone hydrogen bond lengths and CA-atom positions. The four domains were found to be very similar according to all these criteria and superposition of their CA-atoms yielded r.m.s. distances ranging from 0.36 to 0.72Åfor the six possible comparisons. Large deviations (> 1.0Å) are only seen in the five-residue segments that link adjacent domains and at the N and C termini. Refinement has also allowed critical examination of each of the two unique sugar binding sites, referred to as “primary” and “secondary” sites, in different lattice environments. While the essential tyrosyl side-chain in each of these sites (Y73, Y159) assumes precise orientation for optimum hydrophobic contact with the N-acetyl methyl group of the sugar ligand, side-chains involved in hydrogen bonds (S62, E115; and S148, D29) were found to be relatively flexible and able to adapt their conformation to changes in environment. Ordered water structure present in these binding sites is not completely analogous in the different environments. In addition, intermolecular interactions as well as protein-solvent association, important for dimerization and stabilization of the crystal lattice, have been examined and compared in the independent environments.