title: Targeted codon optimization improves translational fidelity for an Fc fusion protein.
abstract: High levels of translational errors, both truncation and misincorporation in an Fc-fusion protein were observed. Here, we demonstrate the impact of several commercially available codon optimization services, and compare to a targeted strategy. Using the targeted strategy, only codons known to have translational errors are modified. For an Fc-fusion protein expressed in Escherichia coli, the targeted strategy, in combination with appropriate fermentation conditions, virtually eliminated misincorporation (proteins produced with a wrong amino acid sequence), and reduced the level of truncation. The use of full optimization using commercially available strategies reduced the initial errors, but introduced different misincorporations. However, truncation was higher using the targeted strategy than for most of the full optimization strategies. This targeted approach, along with monitoring of translation fidelity and careful attention to fermentation conditions is key to minimizing translational error and ensuring high-quality expression. These findings should be useful for other biopharmaceutical products, as well as any other transgenic constructs where protein quality is important.
authors.affiliation: Process and Product Development, Amgen Inc., Thousand Oaks, California. hutterer@amgen.com
authors.forename: Katariina M
authors.initials: KM
authors.lastname: Hutterer
authors.forename: Zhongqi
authors.initials: Z
authors.lastname: Zhang
authors.forename: Mark Leo
authors.initials: ML
authors.lastname: Michaels
authors.forename: Ed
authors.initials: E
authors.lastname: Belouski
authors.forename: Robert W
authors.initials: RW
authors.lastname: Hong
authors.forename: Bhavana
authors.initials: B
authors.lastname: Shah
authors.forename: Mark
authors.initials: M
authors.lastname: Berge
authors.forename: Hedieh
authors.initials: H
authors.lastname: Barkhordarian
authors.forename: Eleanor
authors.initials: E
authors.lastname: Le
authors.forename: Steve
authors.initials: S
authors.lastname: Smith
authors.forename: Dwight
authors.initials: D
authors.lastname: Winters
authors.forename: Frank
authors.initials: F
authors.lastname: Abroson
authors.forename: Randy
authors.initials: R
authors.lastname: Hecht
authors.forename: Jennifer
authors.initials: J
authors.lastname: Liu
mesh_terms: D001709:Biotechnology / Q000379:methods; D003062:Codon; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005285:Fermentation; D007141:Immunoglobulin Fc Fragments / Q000235:genetics* / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D003062:Codon; D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins