Translational repression in vitro by the bacteriophage T4 regA protein

The bacteriophage T4 translational repressor regA protein has been purified from an overproducing strain, and its activity has been studied in simple in vitro protein synthesis reactions. RegA protein was found to inhibit the translation of T4 genes 44, 45, and ORF45-1 in a concentration-dependent fashion. Expression of two other T4 genes which are insensitive to regA protein in vivo, genes 32 and 43, was unaffected by the presence of regA protein. Specific inhibition of synthesis of genes 44, 45, and ORF 45-1 proteins was achieved with 5-20 microM concentrations of regA protein, without the addition of any other T4 encoded proteins or cofactors. When in vitro protein synthesis was performed in two steps, uncoupling translation from transcription, regA protein had an inhibitory effect regardless of whether it was added at the initiation of transcription or only at the translation step. This indicates that regA protein functions during the translation step of protein synthesis in vitro in agreement with previous in vivo studies of regA protein.