title: Cloning and expression of recombinant human GMCSF from Pichia pastoris GS115--a progressive strategy for economic production.
abstract: Human granulocyte-macrophage colony-stimulating factor (hGMCSF) is a proinflammatory cytokine and hematopoietic growth factor. Recombinant human granulocyte-macrophage colony-stimulating factor (rhGMCSF) serves as a biotherapeutic agent in bone marrow stimulations, vaccine development, gene therapy approaches, and stem cell mobilization. The objective of the present study includes construction of rhGMCSF having N-terminal intein tag, expression of protein both extracellularly and intracellularly from yeast expression system followed by its purification in a single step by affinity chromatography. The soluble and biologically active rhGMCSF was obtained from Pichia pastoris GS115. About 122 g DCW/L of final yield was obtained for both cytosolic and secretory expression of Pichia GS115 strain. Purified intracellular hGMCSF was 420 mg/L with a specific activity of 2.1×10⁸ IU/mg, and the purified extracellular recombinant protein was 360 mg/L with a specific activity of 1.9×10⁸ IU/mg. The data presented here indicate the possibilities of exploring the economic ways of producing the rhGMCSF.
authors: 1Centre for Biotechnology, Anna University, Chennai, India; and 2Centre for Bioseparation Technology, VIT University, Vellore, India.
authors: K
authors: K
authors: Srinivasa Babu
authors: Krishna Kanth
authors: KK
authors: Pulicherla
authors: Aju
authors: A
authors: Antony
authors: Sankaranarayanan
authors: S
authors: Meenakshisundaram
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods*; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification; D012995:Solubility
chemical_list: D011993:Recombinant Fusion Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor