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Technical University of Munich

Crystal contact engineering of diverse enzymes to improve technical protein crystallization processes

(M.Sc. Brigitte Walla)

Protein crystallization, an efficient alternative to chromatography in biotechnological downstream processing, is limited so far by the poor crystallizability of many proteins. This thesis demonstrates that rational crystal contact engineering of proteins, particularly introducing electrostatic interactions, significantly enhances the crystallizability of industrial enzymes while preserving enzymatic activity. The approach is scalable and offers a promising route for sustainable bioprocessing.

Publikationen
  • Walla B, Dietrich A-M, Brames E, Bischoff D, Fritzsche S, Castiglione K, Janowski R, Niessing D, Weuster-Botz D (2025): Application of a rational crystal contact engineering strategy on a PET-degrading cutinase. Bioengineering 12: 561.
  • Walla B, Maslakova A, Bischoff D, Janowski R, Niessing D, Weuster-Botz D (2025): Rational introduction of electrostatic interactions at crystal contacts to enhance protein crystallization of an ene reductase. Biomolecules 15: 467.
  • Mentges J, Bischoff D, Walla B, Weuster-Botz D (2024): In-situ microscopy with real-time image analysis enables online monitoring of technical protein crystallization kinetics in stirred crystallizers. Crystals 14: 1009.
  • Bischoff D, Walla B, Wegner CH, Hubbuch J, Weuster-Botz D (2024): Modeling-based monitoring and control of protein crystallization in bioprocesses. In: Kwade A, Kampen I (Eds): Dispersity, structure and phase changes of proteins and bio agglomerates in biotechnological processes. Springer, Cham: 79-117.
  • Wegner CH, Eming SM, Walla B, Bischoff D, Weuster-Botz D, Hubbuch J (2024): Spectroscopic insights into multi-phase protein crystallization in complex lysate using Raman spectroscopy and a particle-free bypass. Front Bioeng Biotech 12: 1397465.
  • Walla B, Bischoff D, Corona Viramontes I, Montes Figueredo S, Weuster-Botz D (2023): Recent advances in the monitoring of protein crystallization processes in downstream processing. Crystals 13: 773.
  • Bischoff D, Walla B, Weuster-Botz D (2022): Machine-learning based protein crystal detection for monitoring of crystallization processes enabled with large-scale synthetic data sets of photorealistic images. Analyt Bioanalyt Chem 414: 6379-6391.
  • Walla B, Bischoff D, Janowski R, von den Eichen N, Niessing D, Weuster-Botz D (2021): Transfer of a rational crystal contact engineering strategy between diverse alcohol dehydrogenases. Crystals11: 975.
  • Grob P, Huber M, Walla B, Hermann J, Janowski R, Niessing D, Hekmat D, Weuster-Botz D (2020) Crystal contact engineering enables efficient capture and purification of an oxidoreductase by technical crystallization. Biotechnol J 15: 2000010.
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