Yeast-based production, purification and bioactivity assay of rainbow trout LECT2
| Autor: | Yu-Hong Shi, Xin-Jiang Lu, Rui-Cheng Zhang, Dan-Yang Yang, Min-Yun Li, Jiong Chen |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Cytotoxicity
Immunologic Fish Proteins endocrine system animal diseases Blotting Western Gene Expression Biology Cathepsin D Pichia law.invention Pichia pastoris Receptors CCR Column chromatography law Gene expression Escherichia coli Animals Respiratory Burst Reverse Transcriptase Polymerase Chain Reaction urogenital system Macrophages Macrophage Activation Chromatography Ion Exchange Head Kidney biology.organism_classification Molecular biology Recombinant Proteins Yeast Respiratory burst Oncorhynchus mykiss Chromatography Gel Recombinant DNA Intercellular Signaling Peptides and Proteins Rainbow trout Target protein |
| Zdroj: | Zoological Research. 34:33 |
| ISSN: | 0254-5853 |
| DOI: | 10.3724/sp.j.1141.2013.01033 |
| Popis: | Leukocyte cell-derived chemotaxin 2 (LECT2) is a secretory cytokine that functions in many physiological and pathological processes. We used a Pichia pastoris expression system for the recombinant expression of rainbow trout LECT2. The recombinant LECT2 was purified by UNOsphere S Cation exchange and size-exclusion chromatography columns. The obtained target protein was highly pure (>96% homogeneity) and the yield was >120 mg/L of yeast cultures. An in vitro chamber assay revealed that recombinant LECT2 could induce chemotactic responses in rainbow trout head kidney-derived macrophages. Recombinant LECT2 not only enhanced macrophage respiratory burst activity and bactericidal activity, but also changed macrophage gene expression. In summary, we established a rapid and efficient method to prepare active recombinant rainbow trout LECT2 using a yeast expression system and column chromatography. Bioactive recombinant LECT2 is essential for studies on protein functions. |
| Databáze: | OpenAIRE |
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