title: Expression, purification and characterization of recombinant human interleukin-2-serum albumin (rhIL-2-HSA) fusion protein in Pichia pastoris.
abstract: Interleukin-2 (IL-2) plays important roles in variety of immune functions. Recombinant IL-2 has become an important therapeutic protein for therapy of melanoma and renal cell carcinoma. Previously, it was proved that the therapeutic efficacy of rIL-2 expressed in Saccharomyces cerevisiae was improved by prolonging its in vivo half-life through genetic fusion with albumin. In this study, a fusion protein composed of hIL-2 genetically fused to HSA was expressed as a secretory protein under AOX1 (alcohol oxidase 1) promoter in Pichia pastoris. An effective strategy was established to express rhIL-2-HSA fusion protein in 5L scale and the optimal purification procedure was investigated. The purity of rhIL-2-HSA in final product was about 95%. The purified rhIL-2-HSA fusion protein could be recognized by both anti-hIL-2 and anti-human serum albumin monoclonal antibody. Bioactivity analysis showed that the purified rhIL-2-HSA fusion protein displayed high level activity on proliferation in IL-2 dependent manner in CTLL2-cells. rhIL-2-HSA fusion protein also showed a extended half-life in plasma compared with IL-2 when tested in a BALB/c mouse model. This study provides an alternative strategy for large-scale production of bioactive rhIL-2-HSA fusion protein using P. pastoris as an expression host.
authors.affiliation: Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, PR China. lyreau@126.com
authors.forename: Jianyong
authors.initials: J
authors.lastname: Lei
authors.forename: Bo
authors.initials: B
authors.lastname: Guan
authors.forename: Bo
authors.initials: B
authors.lastname: Li
authors.forename: Zuoying
authors.initials: Z
authors.lastname: Duan
authors.forename: Yun
authors.initials: Y
authors.lastname: Chen
authors.forename: Huazhong
authors.initials: H
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Jin
mesh_terms: D042931:Aldehyde Oxidase / Q000235:genetics; D000818:Animals; D048708:Cell Growth Processes / Q000502:physiology; D003001:Cloning, Molecular; D005260:Female; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics; D012709:Serum Albumin / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics
chemical_list: D007376:Interleukin-2; D011993:Recombinant Fusion Proteins; D012709:Serum Albumin; D042931:Aldehyde Oxidase