Effects of two cis-acting mutations on the regulation and expression of release factor one in Escherichia coli

Together with release factor (RF) 2, RF1 recognises the stop codons and triggers the hydrolysis of the nascent peptide from peptidyl-tRNA during translation termination. prfA, the gene that codes for RF1, is located at 27 min on the Escherichia coli map as the second gene in the hemA-operon. The concentration of RF1 has been shown to increase with increased growth rate, but it is not known where and how this control is exerted. In this study we show that the growth rate regulation of RF1, at least in part, is controlled at P(hemA1), one of two promoters preceding the hemA gene. We have also characterised two mutations, asuA1 and asuA2, that are antisuppressors to the tRNA suppressor Su2. Our data indicate that the antisuppressor phenotype is caused by an increased amount of RF1. The asuA2 mutation is a G to an A change just downstream of the -10 region of P(hemA1), it leads to a higher concentration of RF1 in the cell and abolishes the growth rate regulation. This indicates that the sequence between the -10 region and the transcription start site is important for growth rate control. The increase in concentration of RF1 caused by asuA1 is most likely at the translational level. The efficiency of translation initiation of prfA is low due to a long distance between the start codon and the Shine-Dalgarno (SD) sequence. The asuA1 mutation creates a new start codon with a more optimal distance to the SD sequence. This leads to an increased expression of RF1, probably due to increased initiation efficiency.