Use of tRNA-Supplemented Host Strains for Expression of Heterologous Genes in E. coli

Abstract

Though widely used, expression of heterologous genes in E. coli can be cumbersome and often fails to yield significant amounts of the desired protein. There are a variety of reasons why a particular heterologous gene fails to yield significant amount of protein, including susceptibility of the protein to degradation or presence of sequences that act as transcriptional pause sites. However, probably the most commonly encountered problem is a mismatch of the codon preference observed in the heterologous gene from the codon usage of the E. coli host. The general problem arises form the fact that due to the redundancy of the genetic code one amino acid can be encoded by more than one codon and different organisms do not utilize each codon at the same frequency (for codon frequencies in E. coli and other organisms, see Table 1). Since E. coli contains 46 different tRNA genes (some of them exist in multiple copies), most codons (but not all) have a corresponding cognate tRNA (1). The relative expression of each tRNA gene is typically matched to the frequency of the corresponding codon in the translated RNA species. Forced, high-level expression of heterologous genes containing codons rarely utilized in E. coli can lead to depletion of the corresponding tRNA pools and subsequently to stalling of the translation process and degradation of the translated mRNA (2–4).