Structural insights into the first step of RNA-dependent cysteine biosynthesis in archaea

Abstract

Cysteine is ligated to tRNA^Cys by cysteinyl-tRNA synthetase in most organisms. However, in methanogenic archaea lacking cysteinyl-tRNA synthetase, O-phosphoserine is ligated to tRNA^Cys by O-phosphoseryl–tRNA synthetase (SepRS), and the phosphoseryl-tRNA^Cys is converted to cysteinyl-tRNA^Cys. In this study, we determined the crystal structure of the SepRS tetramer in complex with tRNA^Cys and O-phosphoserine at 2.6-Å resolution. The catalytic domain of SepRS recognizes the negatively charged side chain of O-phosphoserine at a noncanonical site, using the dipole moment of a conserved α-helix. The unique C-terminal domain specifically recognizes the anticodon GCA of tRNA^Cys. On the basis of the structure, we engineered SepRS to recognize tRNA^Cys mutants with the anticodons UCA and CUA and clarified the anticodon recognition mechanism by crystallography. The mutant SepRS-tRNA pairs may be useful for translational incorporation of O-phosphoserine into proteins in response to the stop codons UGA and UAG.