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New application of depth filters for the immobilization of Candida antarctica lipase B

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Abstract

The objective of this study was to use for the first time depth filters, which are usually intended for clarification of cell culture broth, as a direct immobilization support/matrix for industrially relevant enzymes. With this method, it is not only possible to immobilize pure enzymes; it can be also used for capturing recombinant enzymes directly out of culture supernatant. Therefore, the depth filters were coated with different anionic and cationic polymer layers by Layer-by-Layer (LbL) technology. The immobilization behavior of the model enzyme Candida antarctica lipase B (CalB) was examined. Optimal conditions for lipase immobilization were found for anionic surfaces with Poly (allylamin hydrochlorid) (PAH)/Poly (sodium-4-styrene sulfonate) (PSS) coating in 20 mM acetate buffer pH 4. Stability studies showed that immobilized CalB is 1.7-fold more stable when storage is carried out in buffer at 4 °C, compared to storage in buffer at room temperature or storage after drying at 30 °C for 24 h. The calculated half-life period is 108 days until half of the activity was lost. Furthermore, the possibility of direct capture of the CalB either from sonicated culture broth (Escherichia coli) or from cell-free supernatant was tested. Filter blocking prevented the immobilization of lipase from sonicated culture broth, but immobilization from cell-free supernatant could be performed successfully at moderate biomass content (OD600 = 7.0).

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Acknowledgments

This project is in cooperation between the Institute of Technical Chemistry, Sartorius Stedim Biotech GmbH, and Surflay Nanotec GmbH. It has been carried out as an integral part of the Biocatalysis 2021 Cluster, which is financially supported by the BMBF (Bundesministerium für Bildung und Forschung). Special thanks also to Uwe Bornscheuer and his group from the Institute of Biochemistry, Greifswald University, Germany for providing the expression vector. Funmilola Heinen is gratefully acknowledged for assistance in the laboratory work.

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Authors and Affiliations

  1. Institute of Technical Chemistry, Gottfried Wilhelm Leibniz University of Hannover, Callinstr. 5, 30167, Hannover, Germany

    Sarah Schreiber, Thomas Scheper & Sascha Beutel

  2. Sartorius Stedim Biotech GmbH, August-Spindler-Str. 11, 37079, Göttingen, Germany

    Axel Thiefes

  3. Surflay Nanotec GmbH, Max-Planck-Str. 3, 12489, Berlin, Germany

    Ute Schuldt & Lars Dähne

Authors
  1. Sarah Schreiber
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  2. Axel Thiefes
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  3. Ute Schuldt
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  4. Lars Dähne
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  5. Thomas Scheper
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  6. Sascha Beutel
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Correspondence to Sascha Beutel.

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Schreiber, S., Thiefes, A., Schuldt, U. et al. New application of depth filters for the immobilization of Candida antarctica lipase B. Appl Microbiol Biotechnol 101, 599–607 (2017). https://doi.org/10.1007/s00253-016-7764-5

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  • DOI: https://doi.org/10.1007/s00253-016-7764-5

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