title: GeneOptimizer program-assisted cDNA reengineering enhances sRAGE autologous expression in Chinese hamster ovary cells.
abstract: Soluble receptor for advanced glycation end products (sRAGE) is a secreted mammalian protein that functions as a decoy to counter-react RAGE signaling-resultant pathological conditions, and has high therapeutic potentials. Our prior studies showed that recombinant human sRAGE expressed in Chinese hamster, Ceanothus griseus, ovary (CHO) cells is modified by specific N-glycosylation, and exhibits higher bioactivity than that expressed in other host systems including insect Spodoptera frugiperda cells. Here, we show that GeneOptimizer software program-assisted, reengineered sRAGE cDNA enhances the recombinant protein expression in CHO cells. The cDNA sequence encoding human sRAGE was optimized for RNA structure, stability, and codon usages in CHO cells. We found that such optimization augmented sRAGE expression over 2 folds of its wild-type counterpart. We also studied how individual parameter impacted sRAGE autologous expression in CHO cells, and whether sRAGE bioactivity was compromised. We found that the enhanced expression appeared not to affect sRAGE N-glycosylation and bioactivity. Optimization of sRAGE expression provides a basis for future large-scale production of this protein to meet medical needs.
authors.affiliation: Laboratory of Cardiovascular Sciences, National Institute on Aging, NIH, 251 Bayview Boulevard, Baltimore, MD 21224, United States.
authors.forename: Wen
authors.initials: W
authors.lastname: Wei
authors.affiliation: Laboratory of Cardiovascular Sciences, National Institute on Aging, NIH, 251 Bayview Boulevard, Baltimore, MD 21224, United States.
authors.forename: Ji Min
authors.initials: JM
authors.lastname: Kim
authors.affiliation: GeneArt Division, Life Technologies-Invitrogen Inc., 5823 Newton Drive, Carlsbad, CA 92008, United States.
authors.forename: Danny
authors.initials: D
authors.lastname: Medina
authors.affiliation: Laboratory of Cardiovascular Sciences, National Institute on Aging, NIH, 251 Bayview Boulevard, Baltimore, MD 21224, United States.
authors.forename: Edward G
authors.initials: EG
authors.lastname: Lakatta
authors.affiliation: Laboratory of Cardiovascular Sciences, National Institute on Aging, NIH, 251 Bayview Boulevard, Baltimore, MD 21224, United States. Electronic address: linli@mail.nih.gov.
authors.forename: Li
authors.initials: L
authors.lastname: Lin
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D018076:DNA, Complementary / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D015202:Protein Engineering / Q000379:methods*; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; D011994:Recombinant Proteins
keywords: CHO; GeneOptimizer program; Protein expression; sRAGE