IMPAIRED INVITRO KINETICS OF EF-TU MUTANT AA

The kirromycin‐resistant EF‐Tu mutant Aa, previously shown to be an antisuppressor for nonsense and missense suppressor tRNAs, has been characterised in a poly(U)‐primed translation system in vitro. Two major defects were found in the function of the mutant. First, the dissociation constant for Aa binding to Phe‐tRNAPhe was increased tenfold compared to wild‐type EF‐Tu. Second, kcat/Km for the interaction between the EF‐Tu · GTP · aa‐tRNA complex and the ribosome was decreased by the mutation to one third of its wild‐type value. No differences were observed between mutant and wild‐type factor in the regeneration of EF‐Tu · GTP from EF‐Tu · GDP via EF‐Ts or in the mistranslation frequency by Leu‐tRNALeu4. The relation between the in vitro results and the mutant phenotype in vivo is discussed. Copyright © 1990, Wiley Blackwell. All rights reserved