title: Production of functional human insulin-like growth factor binding proteins (IGFBPs) using recombinant expression in HEK293 cells.
abstract: Insulin-like growth factor binding proteins (IGFBPs) display many functions in humans including regulation of the insulin-like growth factor (IGF) signaling pathway. The various roles of human IGFBPs make them attractive protein candidates in drug discovery. Structural and functional knowledge on human proteins with therapeutic relevance is needed to design and process the next generation of protein therapeutics. In order to conduct structural and functional investigations large quantities of recombinant proteins are needed. However, finding a suitable recombinant production system for proteins such as full-length human IGFBPs, still remains a challenge. Here we present a mammalian HEK293 expression method suitable for over-expression of secretory full-length human IGFBP-1 to -7. Protein purification of full-length human IGFBP-1, -2, -3 and -5 was conducted using a two-step chromatography procedure and the final protein yields were between 1 and 12mg protein per liter culture media. The recombinant IGFBPs contained PTMs and exhibited high-affinity interactions with their natural ligands IGF-1 and IGF-2.
authors.affiliation: Protein Function and Interactions Group, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark. Electronic address: wanscher@cpr.ku.dk.
authors.forename: Anne Sofie Molsted
authors.initials: ASM
authors.lastname: Wanscher
authors.affiliation: Protein Production and Characterization Platform, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark.
authors.forename: Michael
authors.initials: M
authors.lastname: Williamson
authors.affiliation: Protein Production and Characterization Platform, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark.
authors.forename: Tasja Wainani
authors.initials: TW
authors.lastname: Ebersole
authors.affiliation: Protein Function and Interactions Group, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark; Novozymes A/S, Bagsværd, Denmark.
authors.forename: Werner
authors.initials: W
authors.lastname: Streicher
authors.affiliation: Protein Function and Interactions Group, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark.
authors.forename: Mats
authors.initials: M
authors.lastname: Wikström
authors.affiliation: Protein Production and Characterization Platform, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Denmark.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Cazzamali
mesh_terms: D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D018969:Insulin-Like Growth Factor Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D007334:Insulin-Like Growth Factor I / Q000737:chemistry; D007335:Insulin-Like Growth Factor II / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
chemical_list: C583557:IGF1 protein, human; C035170:IGF2 protein, human; D018969:Insulin-Like Growth Factor Binding Proteins; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II
keywords: HEK293 cells; Post-translational modification; Recombinant full-length human IGFBP; Secretory expression; Surface plasmon resonance