title: Efficient and inexpensive method for purification of heparin binding proteins.
abstract: Heparin binding (HB) proteins mediate a wide range of important cellular processes, which makes this class of proteins biopharmaceutically important. Engineering HB proteins may bring many advantages, but it necessitates cost effective and efficient purification methodologies compared to currently available methods. One of the most important classes of HB proteins are fibroblast growth factors (FGFs) and their receptors (FGFRs). In this study, we report an efficient off-column purification of FGF-1 from soluble fractions and purification of the D2 domain of FGFR from insoluble inclusion bodies, using a weak Amberlite cation (IRC) exchanger. FGF-1 and the D2 domain have been expressed in Escherichia coli and purified to homogeneity using IRC resin. This approach is an alternative to conventional affinity column chromatography, which exhibits several disadvantages, including time-consuming experimental procedures for purification and regeneration and results in the expensive production of recombinant proteins. Results of the heparin binding chromatography and steady state fluorescence experiments show that the FGF-1 and the D2 are in a native conformation. The findings of this study will not only aid an in-depth investigation of this class of proteins but will also provide avenues for inexpensive and efficient purification of other important biological macromolecules.
authors.affiliation: Department of Chemistry, Western Kentucky University, Bowling Green, KY 42101, United States.
authors.forename: Sumit
authors.initials: S
authors.lastname: Batra
authors.forename: Nilesh
authors.initials: N
authors.lastname: Sahi
authors.forename: Kristen
authors.initials: K
authors.lastname: Mikulcik
authors.forename: Heather
authors.initials: H
authors.lastname: Shockley
authors.forename: Camille
authors.initials: C
authors.lastname: Turner
authors.forename: Zachary
authors.initials: Z
authors.lastname: Laux
authors.forename: Vivek D
authors.initials: VD
authors.lastname: Badwaik
authors.forename: Eric
authors.initials: E
authors.lastname: Conte
authors.forename: Dakshinamurthy
authors.initials: D
authors.lastname: Rajalingam
mesh_terms: D002852:Chromatography, Ion Exchange / Q000191:economics / Q000379:methods*; D016220:Fibroblast Growth Factor 1 / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry*; D006801:Humans; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017468:Receptors, Fibroblast Growth Factor / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D017468:Receptors, Fibroblast Growth Factor; D011994:Recombinant Proteins; D016220:Fibroblast Growth Factor 1; D006493:Heparin