Recombinant silk protein condensates show widely different properties depending on the sample background

Abstract

There is an increasing understanding that condensation is a crucial intermediate step in the assembly of biological materials and for a multitude of cellular processes. To apply and to understand these mechanisms, in vitro biophysical characterisation techniques are central. The formation and biophysical properties of protein condensates depend on a multitude of factors, such as protein concentration, pH, temperature, salt concentration, and presence of other biomolecules as well as protein purification and storage conditions. Here we show how critical the procedures for preparing protein samples for in vitro studies are. We compare two purification methods of the recombinant spider silk protein CBM-AQ12-CBM and study the effect of background molecules, such as DNA, on the formation and properties of the condensates. We characterize the condensates using aggregation induced emitters (AIEs), coalescence studies, and micropipette aspiration. The condensated sample containing background molecules exhibit a lower threshold concentration for condensate formation accompanied by a lower surface tension and longer coalescence time when compared to the pure protein condensates. Furthermore, the partitioning of small AIEs is enhanced in the presence of background molecules. Our results highlight that the purification method and remaining background molecules strongly affect the biophysical properties of spider silk condensates. Using the acquired knowledge about spider silk protein purification we derive guidelines for reproducible condensate formation that will foster the use of spider silk proteins as adhesives or carriers for biomedical applications.

Graphical abstract: Recombinant silk protein condensates show widely different properties depending on the sample background

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Article information

Article type
Paper
Submitted
29 Jun 2024
Accepted
13 Oct 2024
First published
16 Oct 2024
This article is Open Access
Creative Commons BY license

J. Mater. Chem. B, 2024, Advance Article

Recombinant silk protein condensates show widely different properties depending on the sample background

J. Tersteegen, I. Tunn, M. Sand, T. Välisalmi, M. Malkamäki, J. Gandier, G. Beaune, A. Sanz-Velasco, E. Anaya-Plaza and M. B. Linder, J. Mater. Chem. B, 2024, Advance Article , DOI: 10.1039/D4TB01422G

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