Kinetic studies of constitutive human granulocyte-macrophage colony stimulating factor (hGM-CSF) expression in continuous culture of <Emphasis Type="Italic">Pichia pastoris</Emphasis> |
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Authors: | Yogender Pal Khasa Amardeep Khushoo Leena Srivastava K J Mukherjee |
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Institution: | (1) School of Biotechnology, Jawaharlal Nehru University, New Delhi, 110067, India |
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Abstract: | Extracellular human granulocyte-macrophage colony stimulating factor (hGM-CSF) expression was studied under the control of
the GAP promoter in recombinant Pichia pastoris in a series of continuous culture runs (dilution rates from 0.025 to 0.2 h−1). The inlet feed concentration was also varied and the steady state biomass concentration increased proportionally demonstrating
efficient substrate utilization and constancy of the biomass yield coefficient (Yx/s) for a given dilution rate. The specific product formation rate (qP) showed a strong correlation with dilution rates demonstrating growth associated product formation of hGM-CSF. The volumetric
product concentration achieved at the highest feed concentration (4×) and a dilution rate of 0.2 h−1 was 82 mg l−1 which was 5-fold higher compared to the continuous culture run with 1× feed concentration at the lowest dilution rate thus
translating to a 40 fold increase in the volumetric productivity. The specific product yield (YP/X) increased slightly from 2 to 2.5 mg g−1, with increasing dilution rates, while it remained fairly invariant, for all feed concentrations demonstrating negligible
product degradation or feed back inhibition. The robust nature of this expression system would make it easily amenable to
scale up for industrial production. |
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Keywords: | Constitutive expression Continuous culture GAP Promoter Human granulocyte-macrophage colony stimulating factor Pichia pastoris |
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