title: Tuning of Recombinant Protein Expression in Escherichia coli by Manipulating Transcription, Translation Initiation Rates, and Incorporation of Noncanonical Amino Acids.
abstract: Protein synthesis in cells has been thoroughly investigated and characterized over the past 60 years. However, some fundamental issues remain unresolved, including the reasons for genetic code redundancy and codon bias. In this study, we changed the kinetics of the Eschrichia coli transcription and translation processes by mutating the promoter and ribosome binding domains and by using genetic code expansion. The results expose a counterintuitive phenomenon, whereby an increase in the initiation rates of transcription and translation lead to a decrease in protein expression. This effect can be rescued by introducing slow translating codons into the beginning of the gene, by shortening gene length or by reducing initiation rates. On the basis of the results, we developed a biophysical model, which suggests that the density of co-transcriptional-translation plays a role in bacterial protein synthesis. These findings indicate how cells use codon bias to tune translation speed and protein synthesis.
authors.affiliation: Department of Life Sciences and Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev , PO Box 653, Beer-Sheva 8410501, Israel.
authors.forename: Orr
authors.identifier: 0000-0002-2648-6235
authors.initials: O
authors.lastname: Schlesinger
authors.affiliation: Department of Life Sciences and Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev , PO Box 653, Beer-Sheva 8410501, Israel.
authors.forename: Yonatan
authors.initials: Y
authors.lastname: Chemla
authors.affiliation: Niels Bohr Institute, University of Copenhagen , Blegdamsvej 17, 2100 Copenhagen, Denmark.
authors.forename: Mathias
authors.initials: M
authors.lastname: Heltberg
authors.affiliation: Department of Life Sciences and Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev , PO Box 653, Beer-Sheva 8410501, Israel.
authors.forename: Eden
authors.initials: E
authors.lastname: Ozer
authors.affiliation: School of Physics and Astronomy, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors.forename: Ryan
authors.initials: R
authors.lastname: Marshall
authors.affiliation: School of Physics and Astronomy, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors.forename: Vincent
authors.initials: V
authors.lastname: Noireaux
authors.affiliation: Niels Bohr Institute, University of Copenhagen , Blegdamsvej 17, 2100 Copenhagen, Denmark.
authors.forename: Mogens Høgh
authors.initials: MH
authors.lastname: Jensen
authors.affiliation: Department of Life Sciences and Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev , PO Box 653, Beer-Sheva 8410501, Israel.
authors.forename: Lital
authors.identifier: 0000-0002-3805-8625
authors.initials: L
authors.lastname: Alfonta
mesh_terms: D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D003062:Codon / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008957:Models, Genetic; D014176:Protein Biosynthesis / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D000596:Amino Acids; D003062:Codon; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins
keywords: codon bias; genetic code expansion; protein translation initiation; rates of translation; transcription initiation