title: Simplified in vitro refolding and purification of recombinant human granulocyte colony stimulating factor using protein folding cation exchange chromatography.
abstract: Protein folding-strong cation exchange chromatography (PF-SCX) has been employed for efficient refolding with simultaneous purification of recombinant human granulocyte colony stimulating factor (rhG-CSF). To acquire a soluble form of renatured and purified rhG-CSF, various chromatographic conditions, including the mobile phase composition and pH was evaluated. Additionally, the effects of additives such as urea, amino acids, polyols, sugars, oxidizing agents and their amalgamations were also investigated. Under the optimal conditions, rhG-CSF was efficaciously solubilized, refolded and simultaneously purified by SCX in a single step. The experimental results using ribose (2.0M) and arginine (0.6M) combination were found to be satisfactory with mass yield, purity and specific activity of 71%, ≥99% and 2.6×10(8)IU/mg respectively. Through this investigation, we concluded that the SCX refolding method was more efficient than conventional methods which has immense potential for the large-scale production of purified rhG-CSF.
authors.affiliation: K L E F University, Centre for Bioprocess Technology, Department of Biotechnology, Guntur 522 502, Andhra Pradesh, India.
authors.forename: Sandeep
authors.initials: S
authors.lastname: Vemula
authors.affiliation: K L E F University, Centre for Bioprocess Technology, Department of Biotechnology, Guntur 522 502, Andhra Pradesh, India.
authors.forename: Akshay
authors.initials: A
authors.lastname: Dedaniya
authors.affiliation: K L E F University, Centre for Bioprocess Technology, Department of Biotechnology, Guntur 522 502, Andhra Pradesh, India.
authors.forename: Rahul
authors.initials: R
authors.lastname: Thunuguntla
authors.affiliation: K L E F University, Centre for Bioprocess Technology, Department of Biotechnology, Guntur 522 502, Andhra Pradesh, India.
authors.forename: Maheswara Reddy
authors.initials: MR
authors.lastname: Mallu
authors.affiliation: Andhra University, College of Engineering, Visakhapatnam 530 003, Andhra Pradesh, India.
authors.forename: Pavani
authors.initials: P
authors.lastname: Parupudi
authors.affiliation: K L E F University, Centre for Bioprocess Technology, Department of Biotechnology, Guntur 522 502, Andhra Pradesh, India. Electronic address: rsr@kluniversity.in.
authors.forename: Srinivasa Reddy
authors.initials: SR
authors.lastname: Ronda
mesh_terms: D002412:Cations / Q000737:chemistry; D002845:Chromatography; D004926:Escherichia coli / Q000737:chemistry; D016179:Granulocyte Colony-Stimulating Factor / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D066298:In Vitro Techniques; D008970:Molecular Weight; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry*; D014508:Urea / Q000737:chemistry
chemical_list: D002412:Cations; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D014508:Urea
keywords: Cation exchange chromatography; Mass yield; PFLC; Purity; Specific activity