Mutations replacing the leucine codons or altering the length of the amino acid-coding portion of the ilvGMEDA leader region of Escherichia coli

Abstract

The specificity of regulation by attenuation of the ilvGMEDA operon of Escherichia coli was examined by making alterations in the peptide-coding portion of the leader region. The effects of the alterations on attenuation control were monitored by operon fusions with the lacZ or cat gene. Substitution of the tandem leucine codons with arginine codons did not result in arginine control of attenuation even though the altered leader transcripts contained three consecutive arginine codons. Substitution of the single leucine codon with a proline codon at position 10 of the putative peptide, which had been shown to be important in the regulation of the Serratia marcescens ilv operon, did not result in control of attenuation by proline. Since the formation of neither proline nor arginine biosynthetic enzymes is regulated by attenuation control, the effect of tandem phenylalanine codons in place of the tandem leucine codons was examined and found not to result in control by phenylalanine supply. The latter failure may have been due to a configuration in the secondary structure of the protector stem of the leader transcript different from that of the wild-type transcript. The results of the study favored the idea that the lead ribosome does not initiate translation of the leader transcript until after the RNA polymerase has reached the pause site (117 bases into the leader region).