title: Effect of N-terminal solubility enhancing fusion proteins on yield of purified target protein.
abstract: We have studied the effect of solubilising N-terminal fusion proteins on the yield of target protein after removal of the fusion partner and subsequent purification using immobilised metal ion affinity chromatography. We compared the yield of 45 human proteins produced from four different expression vectors: three having an N-terminal solubilising fusion protein (the GB1-domain, thioredoxin, or glutathione S-transferase) followed by a protease cleavage site and a His tag, and one vector having only an N-terminal His tag. We have previously observed a positive effect on solubility for proteins produced as fusion proteins compared to proteins produced with only a His tag in Escherichia coli. We find this effect to be less pronounced when we compare the yields of purified target protein after removal of the solubilising fusion although large target-dependent variations are seen. On average, the GB1+His fusion gives significantly higher final yields of protein than the thioredoxin+His fusion or the His tag, whereas GST+His gives lower yields. We also note a strong correlation between solubility and target protein size, and a correlation between solubility and the presence of peptide fragments that are predicted to be natively disordered.
authors.affiliation: Department of Biotechnology, Royal Institute of Technology (KTH), SE-106 91, Stockholm, Sweden.
authors.forename: Martin
authors.initials: M
authors.lastname: Hammarström
authors.forename: Esmeralda A
authors.initials: EA
authors.lastname: Woestenenk
authors.forename: Niklas
authors.initials: N
authors.lastname: Hellgren
authors.forename: Torleif
authors.initials: T
authors.lastname: Härd
authors.forename: Helena
authors.initials: H
authors.lastname: Berglund
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D012995:Solubility
chemical_list: D011993:Recombinant Fusion Proteins