Kinetics and active fraction determination of a protease enzyme immobilized on functionalized membranes: mathematical modeling and experimental results.
A detailed study on the performance of a membrane bioreactor is presented, considering diffusion reaction models with product adsorption and structure-function correlations. The enzyme papain was utilized for experimental investigation both in the homogeneous state and on a modified polysulfone (MPS) membrane. Variation of enzyme loading on the membrane and enzyme concentration in the solution state depicted decreasing activity with increasing loading. The performance of the bioreactor was simulated using a diffusion reaction model within a recirculation loop. Electron paramagnetic resonance (EPR) spectroscopy was utilized to study the conformational changes of the active site of papain immobilized on the MPS membrane. Two models were applied to correlate the structure and function of the biocatalyst, based on loading (kinetics) and EPR (structure). The active fractions, lambda, determined from the two models were 0.29 and in the range 0.25-0.3, respectively. The intrinsic kinetics (Vmax) for the immobilized enzyme as determined by the correlations were in the range 101-121 micromol/(g.min), compared to 111 micromol/(g.min) for the homogeneous enzyme. This proves that the immobilized enzyme kinetics do approach homogeneous kinetics for papain on the MPS membrane, when corrected for adsorption and conformational changes.
Ganapathi-Desai
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Department of Chemical and Materials Engineering, Department of Chemistry, and Center of Membrane Sciences, University of Kentucky, Lexington, Kentucky 40506-0046, USA.
Butterfield
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DA
Bhattacharyya
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D002384:Catalysis; D020134:Catalytic Domain; D004578:Electron Spin Resonance Spectroscopy; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008567:Membranes, Artificial; D008962:Models, Theoretical; D010206:Papain / Q000737:chemistry* / Q000378:metabolism*; D011108:Polymers; D011487:Protein Conformation; D013450:Sulfones
D004800:Enzymes, Immobilized; D008567:Membranes, Artificial; D011108:Polymers; D013450:Sulfones; C017662:polysulfone P 1700; D010206:Papain