Abstract
A complex formed between the dimeric aspartyl-tRNA synthetase from yeast (Mr congruent to 125,000) and two molecules of its cognate yeast tRNAAsp (Mr = 24,160) was crystallized using ammonium sulfate as the precipitant. The crucial parameter which governs a successful crystallization is the enzyme tRNA stoichiometry. Crystals are only obtained when the starting solution precisely contains two tRNA molecules for one enzyme molecule. It was demonstrated by electrophoresis, biological activity assays, and crystallographic data that the crystals contain the two components in the same two to one stoichiometric ratio. The crystals, of cubic shape with edges up to 0.8 mm, belong to space group 1432. The cell parameter is 354 A and the asymmetric unit contains one particle of complex. The solvent content is about 78%, higher than the values commonly observed. Although particularly soft, the quality of the crystals is suitable for x-ray diffraction studies up to 7-A resolution.
Highlights
A complex formed betweenthe dimeric aspartyltRNA synthetase from yeast (Mr= 125,000) and two molecules of its cognate yeast tRNAAsP(Mr= 24,160)
High ionicstrength was thought to hampceormplex formation, it was shown that the aminoacylatiorneaction could occur at high ammonium sulfate concentrations (14)
As for thespherical viruses,inall the structures The successful crystallization of the complex formed besolved a t highresolution the RNA part is disordered and tween tRNAAspand aspartyl-tRNA synthetase using ammocannot be seen on the electron density map(3s8-40)
Summary
A complex formed betweenthe dimeric aspartyltRNA synthetase from yeast (Mr= 125,000) and two molecules of its cognate yeast tRNAAsP(Mr= 24,160). Crystals are only obtained when the starting solution precisely contains twotRNA molecules for one enzyme molecule. It was demonstrated by electrophoresis, biological activity assays,and crystallographic data that the crystals contain the two components in the same two to one stoichiometric ratio. After the first successful crystallization of yeast tRNAAsP (12), the focus was on the asparticacid system from yeast and suitablecrystals of thecognateaspartyl-tRNAsynthetase were soon obtained using ammonium sulfate as the precipitant (13). High ionicstrength was thought to hampceormplex formation, it was shown that the aminoacylatiorneaction could occur at high ammonium sulfate concentrations (14). After this positive result crystallization attempts were successfully carried on. Preliminary and partial results of this work have been published as a short communication(15)
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