title: Quantifying translational coupling in E. coli synthetic operons using RBS modulation and fluorescent reporters.
abstract: Translational coupling is the interdependence of translation efficiency of neighboring genes encoded within an operon. The degree of coupling may be quantified by measuring how the translation rate of a gene is modulated by the translation rate of its upstream gene. Translational coupling was observed in prokaryotic operons several decades ago, but the quantitative range of modulation translational coupling leads to and the factors governing this modulation were only partially characterized. In this study, we systematically quantify and characterize translational coupling in E. coli synthetic operons using a library of plasmids carrying fluorescent reporter genes that are controlled by a set of different ribosome binding site (RBS) sequences. The downstream gene expression level is found to be enhanced by the upstream gene expression via translational coupling with the enhancement level varying from almost no coupling to over 10-fold depending on the upstream gene's sequence. Additionally, we find that the level of translational coupling in our system is similar between the second and third locations in the operon. The coupling depends on the distance between the stop codon of the upstream gene and the start codon of the downstream gene. This study is the first to systematically and quantitatively characterize translational coupling in a synthetic E. coli operon. Our analysis will be useful in accurate manipulation of gene expression in synthetic biology and serves as a step toward understanding the mechanisms involved in translational expression modulation.
authors: Department of Plant Sciences, Weizmann Institute of Science , Rehovot 76100, Israel.
authors: Ayelet
authors: A
authors: Levin-Karp
authors: Uri
authors: U
authors: Barenholz
authors: Tasneem
authors: T
authors: Bareia
authors: Michal
authors: M
authors: Dayagi
authors: Lior
authors: L
authors: Zelcbuch
authors: Niv
authors: N
authors: Antonovsky
authors: Elad
authors: E
authors: Noor
authors: Ron
authors: R
authors: Milo
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D009876:Operon / Q000235:genetics*; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis; D060888:Real-Time Polymerase Chain Reaction; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D008164:Luminescent Proteins; C054562:yellow fluorescent protein, Bacteria