N‐Acetyl lysyl‐tRNA synthetases evolved by a CcdB‐based selection possess N‐acetyl lysine specificity in vitro and in vivo

摘要

Posttranslationalmodificationsplayacrucialroleinmodulatingproteinstructureandfunction.Geneticincorporationofunnaturalaminoacidsintoaspecificsiteofaproteinfacilitatesthesystematicstudyofproteinmodificationsincludingacetylation.Weherereportthedirectedevolutionofpyrrolysyl‐tRNAsynthetase(PylRS)fromMethanosarcinamazeitocreateN‐acetyllysyl‐tRNAsynthetases(AcKRSs)usinganewselectionsystembasedonthekillingactivityofthetoxicccdBgeneproduct.Theaminoacidspecificityoftheseandofpublished[1,2]AckRSswastestedinvitroandinvivo,andtheenzyme‐kineticpropertiesoftheAckRSswereevaluatedforthefirsttime...