Optimized Removal of Soluble Host Cell Proteins for the Recovery of met-Human Growth Hormone Inclusion Bodies from Escherichia coli Cell Lysate Using Crossflow Microfiltration

Authors

  • Adith Venkiteshwaran,

    1. Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York
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  • Patrick Heider,

    1. Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York
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  • Sandro Matosevic,

    1. Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York
    Current affiliation:
    1. Department of Biochemical Engineering, University College London, London WC1E 7 JE, U.K.
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  • Are Bogsnes,

    1. Novo Nordisk A/S, Protein Separation, Gentofte, Denmark
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  • Arne Staby,

    1. Novo Nordisk A/S, Protein Separation, Gentofte, Denmark
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  • Susan Sharfstein,

    1. Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York
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  • Georges Belfort

    Corresponding author
    1. Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York
    • Howard P. Isermann Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York. Tel: 518–276–6948.
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Abstract

Cross-flow membrane microfiltration was used under optimal conditions to recover met-growth hormone inclusion bodies (IBs) from Escherichia coli cell lysate by removal of the host-cell (bacterial) proteins (HCP) under minimal fouling conditions. This is the first step of a two-step process in which the goal was to isolate IBs at high yield from the HCP. These undesired soluble HCP were removed by passing them through the membrane while retaining the insolubles, including the aggregated IBs. Experiments were conducted at constant permeate flux with flat-sheet membranes of different pore sizes and chemistry, with feeds of varying pH and ionic strengths to determine the optimum combination for HCP removal. Diafiltration, the washing away of impurities with protein-free buffer, was then employed to ensure removal of the host cell proteins at the optimum conditions. About 90% removal of the HCP was obtained in about 5 diavolumes, maintaining high protein transmission and low membrane fouling.

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