title: Portable Shine-Dalgarno regions; nucleotides between the Shine-Dalgarno sequence and the start codon affect the translation efficiency.
abstract: This chapter describes a gene expression system of E. coli that contains a portable Shine-Dalgarno region. Transcription of this system is directed by a hybrid promoter derived from trp and lac-UV5 promoter sequences, which is followed by a region that encodes the portable Shine-Dalgarno region. We used a series of synthetic portable Shine-Dalgarno regions to vary the length (from 4 to 13 bases) of the Shine-Dalgarno by increasing the number of bases on the mRNA that are complementary to the 3' end of 16s rRNA. Increase in the Shine-Dalgarno region to 8 or 13 bases decreased the translation efficiency of the chimeric leukocyte interferon messenger by 40%. In another series of portable Shine-Dalgarno regions, we varied the four bases that follow the Shine-Dalgarno region. The presence of four A residues or four T residues in this region resulted in the highest translation efficiency. The presence of four C residues reduced translation efficiency by 50%, compared with A or T residues. The presence of four G residues after the Shine-Dalgarno region lowered translation efficiency by 75%, compared with A or T residues.
mesh_terms: D003001:Cloning, Molecular; D004247:DNA / Q000138:chemical synthesis; D004926:Escherichia coli / Q000235:genetics*; D013006:Growth Hormone / Q000096:biosynthesis; D007372:Interferons / Q000096:biosynthesis; D010442:Peptide Chain Initiation, Translational; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D012689:Sequence Homology, Nucleic Acid
chemical_list: D012329:RNA, Bacterial; D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S; D013006:Growth Hormone; D004247:DNA; D007372:Interferons