Integration of mechanical cell disruption and fluidised bed recovery of G3PDH from unclarified disrupted yeast: a comparative study of the performance of unshielded and polymer shielded dye-ligand chromatography systems
The development of a simplified process for the simultaneous disruption and direct selective purification of intracellular proteins from unclarified yeast disruptate has been investigated. The recovery of glyceraldehyde 3-phosphate dehydrogenase (G3PDH) from baker's yeast was selected as a pote...
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| Main Authors: | , |
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| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2005
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| Online Access: | http://psasir.upm.edu.my/id/eprint/15488/1/Integration%20of%20mechanical%20cell%20disruption%20and%20fluidised%20bed%20recovery%20of%20G3PDH%20from%20unclarified%20disrupted%20yeast%20a%20comparative%20study%20of%20the%20performance%20of%20unshielded%20and%20polymer%20shielded%20dye-ligand%20chromatography%20systems.pdf http://psasir.upm.edu.my/id/eprint/15488/ https://www.sciencedirect.com/science/article/pii/S0168165605002944#! |
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| Summary: | The development of a simplified process for the simultaneous disruption and direct selective purification of intracellular proteins from unclarified yeast disruptate has been investigated. The recovery of glyceraldehyde 3-phosphate dehydrogenase (G3PDH) from baker's yeast was selected as a potential demonstration of the generic applicability and practical feasibility of this integrated technique. The application of an adsorbent characterised by high density (UpFront steel-agarose; ρ = 2.65 g ml−1) facilitated the combining of cell disruption operation (bead milling of 50% ww/v of yeast suspension at 7.2 l h−1) with fluidised bed dye-ligand (Cibacron Blue 3GA) adsorption operated immediately downstream of the disrupter. The adoption of a polymer shielded, dye-ligand technique advanced recovery efficiency. It was demonstrated that G3PDH could be recovered with a yield of 67.5% bound activity and a specific activity of 40.2 IU mg−1, after a single step elution with 0.15 M NaCl. The generic application of this approach has been evaluated. |
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