Engineered Aminoacyl-tRNA Synthetase for Cell-Selective Analysis of Mammalian Protein Synthesis

Methods
for cell-selective analysis of proteome dynamics will facilitate
studies of biological processes in multicellular organisms. Here we
describe a mutant murine methionyl-tRNA synthetase (designated L274GMmMetRS) that charges the noncanonical amino acid azidonorleucine
(Anl) to elongator tRNAMet in hamster (CHO), monkey (COS7),
and human (HeLa) cell lines. Proteins made in cells that express the
synthetase can be labeled with Anl, tagged with dyes or affinity reagents,
and enriched on affinity resin to facilitate identification by mass
spectrometry. The method does not require expression of orthogonal
tRNAs or depletion of canonical amino acids. Successful labeling of
proteins with Anl in several mammalian cell lines demonstrates the
utility of L274GMmMetRS as a tool for cell-selective
analysis of mammalian protein synthesis.