The frequency of transcriptional and translational errors at nonsense codons in the lacZ gene of Escherichia coli

Abstract

Nonsense alleles in the lacZ gene of E. coli do not completely eliminate enzyme activity as errors during protein synthesis allow some chains to be completed. The relative contributions of transcriptional and translational errors to this leakiness were investigated by two methods: the introduction of rho- alleles into extreme-polar mutants and the kinetics of beta-galactosidase induction. Virtually all the errors appeared to be transcriptional in the case of two extreme-polar and one non-polar mutation. These alleles should prove useful for further in vivo investigations of RNA polymerase accuracy. With two other non-polar alleles, transcriptional mistakes were low and translational ones high. The frequency of RNA polymerase errors was context-dependent and varied for different nonsense codons in the same position and for the same codon in different positions. The reasons why some alleles showed no activity due to translational errors could not be clearly established. However, increasing the rates of ribosomal errors from one such allele with streptomycin raised the contribution of ribosomal errors to activity markedly and non-linearly. Translational mistakes may give rise to active enzyme only if the monomers are formed at a rate sufficient for effective aggregation to the normal tetramer.