title: High level expression and purification of active recombinant human interleukin-8 in Pichia pastoris.
abstract: Human interleukin-8 (hIL-8) is a member of interleukin family which functions as a chemotactic factor as well as an angiogenesis mediator. Previously, a study reported that hIL-8 could be purified from inclusion bodies using a prokaryotic expression system, however, the required re-naturation step limits the recovery of fully active protein. In this study, soluble recombinant hIL-8 was expressed as a secreted protein at high level in Pichia pastoris under the control of AOX1 (alcohol oxidase 1) promoter. A simple purification strategy was established to recover rhIL-8 from the fermentation supernatant. The process includes precipitation with 80% saturation ammonium sulfate and CM Sepharose ion exchange chromatography, yielding 30 mg/L purified rhIL-8 at over 95% purity. The obtained rhIL-8 displays high specific activity, stimulating the migration of mouse neutrophils at concentrations as low as 0.25 ng/mL. Our results demonstrate that P. pastoris expression system is an efficient tool for large-scale manufacture of active recombinant hIL-8 for various applications.
authors: Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
authors: Hongbo
authors: H
authors: Li
authors: Dongye
authors: D
authors: Wang
authors: Aimin
authors: A
authors: Xu
authors: Shiwu
authors: S
authors: Li
authors: Shouguang
authors: S
authors: Jin
authors: Donghai
authors: D
authors: Wu
mesh_terms: D000645:Ammonium Sulfate / Q000737:chemistry; D000818:Animals; D002465:Cell Movement; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D005285:Fermentation; D006801:Humans; D016209:Interleukin-8 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009504:Neutrophils; D010843:Pichia / Q000378:metabolism*; D010865:Pilot Projects; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012995:Solubility
chemical_list: D016209:Interleukin-8; D011994:Recombinant Proteins; D000645:Ammonium Sulfate