title: The ribosomal exit tunnel as a target for optimizing protein expression in Escherichia coli.
abstract: The folding of many cellular proteins occurs co-translationally immediately outside the ribosome exit tunnel, where ribosomal proteins and other associated factors coordinate the synthesis and folding of newly translated polypeptides. Here, we show that the large subunit protein L29, which forms part of the exit tunnel in Escherichia coli, is required for the productive synthesis of an array of structurally diverse recombinant proteins including the green fluorescent protein (GFP) and an intracellular single-chain Fv antibody. Surprisingly, the corresponding mRNA transcript level of these proteins was markedly less abundant in cells lacking L29, suggesting an unexpected regulatory mechanism that links defects in the exit tunnel to the expression of genetic information. To further highlight the importance of L29 in maintaining protein expression, we used mutagenesis and selection to obtain L29 variants that enhanced GFP expression. Overall, our results suggest that the ribosomal exit tunnel proteins may be key targets for optimizing the overproduction of active, structurally complex recombinant proteins in bacterial cells.
authors.affiliation: School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, NY 14853, USA.
authors.forename: Lydia M
authors.initials: LM
authors.lastname: Contreras-Martinez
authors.forename: Jason T
authors.initials: JT
authors.lastname: Boock
authors.forename: Jan S
authors.initials: JS
authors.lastname: Kostecki
authors.forename: Matthew P
authors.initials: MP
authors.lastname: DeLisa
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D049452:Green Fluorescent Proteins; D008958:Models, Molecular; D017510:Protein Folding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism*; D012270:Ribosomes / Q000235:genetics* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; D049452:Green Fluorescent Proteins
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