Figure 4. Functional switch of LysRS by phospho-Ser207.

(A) Phospho-mimetic LysRS^S207D has enhanced Ap₄A synthetic activity.

(B) Phospho-mimetic LysRS^S207D has enhanced ATP hydrolysis.

(C) Phospho-mimetic LysRS^S207D loses aminoacylation activity for tRNA^Lys. Four concentrations of tRNA were used for this assay. Error bars are standard deviations (SDs) from triplicates.

(D) Phosphorylation of Ser207 redirects the enzymatic flow of LysRS from aminoacylation to Ap₄A synthesis to activate transcription.

(E) Functional replacement assays in yeast show that LysRS^S207D is defective in essential translational functions in cells. The expression of endogenous yeast LysRS can be switched off by the addition of doxycycline to the yeast growth medium. Ten-fold serial dilutions of freshly grown yeast cells were spotted onto selective media SCM-HIS containing 2% raffinose with or without doxycycline and galactose.