Production and purification of recombinant human granulocyte-macrophage colony stimulating factor (GM-CSF) from high cell density cultures of Pichia pastoris.
Granulocyte-macrophage colony stimulating factor (GM-CSF) is a hematopoietic growth factor, which has been used as a therapeutic agent in clinical cases like neutropenia. In this study, we report the production of recombinant human GM-CSF in the methylotrophic yeast Pichia pastoris through secretory expression using the inducible AOX1 promoter. Recombinant P. pastoris GS115 cells were grown in fed batch cultures to obtain a biomass density of 55.6 gDCW L(-1) and a high volumetric activity of 131 mg L(-1) of GM-CSF. The protein migrated as a diffuse band on SDS-PAGE at the range of 28-35 kDa indicating differential glycosylation. The secreted protein was purified to 95% in two steps using cation exchange and size exclusion chromatography.
Bhatacharya
Palash
P
Centre for Biotechnology, Jawaharlal Nehru University, New Delhi, 110067, India.
Pandey
Gaurav
G
Mukherjee
Krishna Jyoti
KJ
D018533:Biomass; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D002412:Cations; D002852:Chromatography, Ion Exchange / Q000379:methods; D005822:Genetic Vectors; D006031:Glycosylation; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000737:chemistry*; D006413:Hematopoietic System / Q000196:embryology; D006801:Humans; D010843:Pichia / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000737:chemistry; D015203:Reproducibility of Results
D002412:Cations; D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor