Crystallographic analysis of a subcomplex of the transsulfursome with tRNA for Cys-tRNA^Cys synthesis

In most organisms, Cys-tRNA^Cys is directly synthesized by cysteinyl-tRNA synthetase (CysRS). Many methanogenic archaea, however, use a two-step, indirect pathway to synthesize Cys-tRNA^Cys owing to a lack of CysRS and cysteine-biosynthesis systems. This reaction is catalyzed by O-phosphoseryl-tRNA synthetase (SepRS), Sep-tRNA:Cys-tRNA synthase (SepCysS) and SepRS/SepCysS pathway enhancer (SepCysE) as the transsulfursome, in which SepCysE connects both SepRS and SepCysS. On the transsulfursome, SepRS first ligates an O-phosphoserine to tRNA^Cys, and the mischarged intermediate Sep-tRNA^Cys is then transferred to SepCysS, where it is further modified to Cys-tRNA^Cys. In this study, a subcomplex of the transsulfursome with tRNA^Cys (SepCysS-SepCysE-tRNA^Cys), which is involved in the second reaction step of the indirect pathway, was constructed and then crystallized. The crystals diffracted X-rays to a resolution of 2.6 Å and belonged to space group P6₅22, with unit-cell parameters a = b = 107.2, c = 551.1 Å. The structure determined by molecular replacement showed that the complex consists of a SepCysS dimer, a SepCysE dimer and one tRNA^Cys in the asymmetric unit.