2.9 Å CRYSTAL STRUCTURE OF LIGAND-FREE TRYPTOPHANYL-TRNA SYNTHETASE Valentin Ilyin, Charles W. Carter, Jr., Evon Winborne, Brenda Temple, and GenPei Li* Department of Biochemistry and Biophysics, CB 7260 University of North Carolina at Chapel Hill, Chapel Hill, NC 27514 USA and *Department of Physical Chemistry, Beijing University, Beijing, Peoples Republic of China

Ligand-free Bacillus stearothermophilus Tryptophanyl-tRNA synthetase (TrpRS) crystallizes either as triclinic (P1) or monoclinic (C2) forms. Systematic response surface analysis of the crystal growth conditions has revealed an optimum at about 42(C, consistent with the thermophilic origin of the enzyme. Unit cell dimensions and 32 noncrystallographic symmetry in both asymmetric units are very similar to those previously observed for monoclinic (P21) crystals of crystals grown in the presence of tryptophan. We have placed the known structure using AMORE, and shown that each asymmetric unit contains either one or two units of three enzyme dimers characterized by 3₁ screw symmetry. Thus, all three crystal forms are closely related to the primitive space group P3₁2₁, previously observed at 18 Å resolution in the P2₁ crystals, and thought to belong to space group P32₁ (Carter, et al., 1990, Acta Cryst. A46:57-68).

We are using isomorphous replacement with selenomethionine-substituted TrpRS (Doublié, et al., 1994, Acta Cryst A50:164-182), together with non-crystallographic symmetry averaging and maximum entropy solvent flattening to supplement phases from positioned fragments of the known model for structure determination. Conformational differences between the different TrpRS structures, including a detailed analysis of the composition and properties of nonpolar nuclei and microclusters by the method of Ilyin (1994 Prot. Eng. 7:1189-1198) will be described.

(This work supported by NIH GM48519-02)