Catalytically Active Inclusion Bodies─Benchmarking and Application in Flow Chemistry

ACS Synthetic Biology, vol. 11, pp. 1881–1896

Abstract

In industries, enzymes are often immobilized to obtain stable preparations that can be utilized in batch and flow processes. In contrast to traditional immobilization methods that rely on carrier binding, various immobilization strategies have been recently presented that enable the simultaneous production and in vivo immobilization of enzymes. Catalytically active inclusion bodies (CatIBs) are a promising example for such in vivo enzyme immobilizates. CatIB formation is commonly induced by fusion of aggregation-inducing tags, and numerous tags, ranging from small synthetic peptides to protein domains or whole proteins, have been successfully used. However, since these systems have been characterized by different groups employing different methods, a direct comparison remains difficult, which prompted us to benchmark different CatIB-formation-inducing tags and fusion strategies. Our study highlights that important CatIB properties like yield, activity, and stability are strongly influenced by tag selection and fusion strategy. Optimization enabled us to obtain alcohol dehydrogenase CatIBs with superior activity and stability, which were subsequently applied for the first time in a flow synthesis approach. Our study highlights the potential of CatIB-based immobilizates, while at the same time demonstrating the robust use of CatIBs in flow chemistry.

Authors 8

1. Gizem Ölçücü

   Forschungszentrum Jülich · Heinrich Heine University Düsseldorf

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

   Institute of Molecular Enzyme Technology, Heinrich Heine University Düsseldorf, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

2. Benedikt Baumer

   Forschungszentrum Jülich · Heinrich Heine University Düsseldorf

   Affiliation as printed

   Institute of Biorganic Chemistry, Heinrich Heine University Düsseldorf, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

3. Kira Küsters

   Forschungszentrum Jülich

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

4. Kathrin Möllenhoff

   Heinrich Heine University Düsseldorf

   Affiliation as printed

   Mathematical Institute, Heinrich Heine University Düsseldorf, Universitätsstrasse 1, 40225 Düsseldorf, Germany

5. Marco Oldiges Aachen Institute of Biotechnology

   Forschungszentrum Jülich · RWTH Aachen University

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

   Institute of Biotechnology, RWTH Aachen University, Worringer Weg 3, D-52074 Aachen, Germany

6. Jörg Pietruszka

   Forschungszentrum Jülich · Heinrich Heine University Düsseldorf

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

   Institute of Biorganic Chemistry, Heinrich Heine University Düsseldorf, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

7. Karl‐Erich Jaeger

   Forschungszentrum Jülich · Heinrich Heine University Düsseldorf

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

   Institute of Molecular Enzyme Technology, Heinrich Heine University Düsseldorf, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

8. Ulrich Krauß corresponding

   Forschungszentrum Jülich · Heinrich Heine University Düsseldorf

   Affiliation as printed

   IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany

   Institute of Molecular Enzyme Technology, Heinrich Heine University Düsseldorf, Forschungszentrum Jülich GmbH, Wilhelm Johnen Strasse, D-52425 Jülich, Germany