5'-Terminal nucleotide sequence of Escherichia coli lactose repressor mRNA: features of translational initiation and reinitiation sites

Abstract

In a sequence of 214 nucleotides at the 5' terminus of the I gene mRNA, which codes for the lactose repressor protein of Escherichia coli, (i) an untranslated leader sequence of 28 residues precedes the repressor coding region; (ii) a GUG initiates synthesis of the wild-type repressor; (iii) GUG and AUG are the functional initiators for the synthesis of restart polypeptides activated by early I gene amber mutations, confirming previous assignments for these residues based on protein sequencing data; and (iv) sequences complementary to 16S ribosomal RNA provide stronger potential mRNA.16S rRNA interaction at the wild-type initiation site than at the restart sites. When I mRNA is used to direct the formation of initiation complexes in vitro, ribosomes bind only to the wild-type initiator region.A striking feature of the I mRNA sequence is the presence of a number of in-phase GUGs that have not been observed to serve as initiation signals in vivo in the nonsense mutant strains examined. The selective use of potential initiator triplets in the I mRNA leads to the following conclusions. First, when presented with several neighboring initiator triplets at the wild-type initiator region, ribosomes select the one preceded by the strongest appropriately positioned complementarity to the 16S 3' end. Second, ribosomes do not restart after termination simply by moving to the next available initiator codon. Third, the formation of stable secondary structures predicted for the untranslated I mRNA beyond chain-terminating nonsense mutations may prevent ribosome access to some potential reinitiation sites.