My project

Proteins from extremophile organisms offer exciting possibilities and huge potential for advanced IB applications. One such rapidly expanding IB sector is that of biopharmaceuticals, where methods to assess the suitability of candidate proteins for bio-processing and knowledge of how to rationally enhance their bio-processability are lacking, leading to significant development lead-time. We propose a new strategy for assessing the potential of protein candidates for bio-processing by uncovering the “design principles” of protein stabilisation [1] inspired by the enhanced stability of proteins from the thermoacidophilic red microalga Galdieria. The project firmly aligns with the theme of “Industrial Biotechnology and Bioenergy” and exploits novel instrumentation for extremophilic protein stability characterization developed using ERC funding. As well as providing fundamental knowledge of the relationship between the stability and solubility of the proteins, the methods developed will allow predictive insights providing an opportunity to design proteins for specific IB needs. Our aim is thus to derive new fundamental knowledge of the stability of extremophilic proteins from Galdieria which have widespread industrial relevance, as well as a practical route towards assessing and improving the quality of protein pharmaceuticals during the production process.

Biopharmaceuticals are currently dominated by proteins derived from antibodies and their simplified derivatives [2]. We will demonstrate and optimise the applicability of our approach using proteins with Ig-like folds, as previously studied by the PI [3]. Using recently developed methods, we will determine the stability and aggregation propensity of Ig-fold proteins from Galdieria. We will mine Galdieria genomes to find “all” Ig-fold proteins and characterise the biotechnological potential of an interesting sub-class (3 genomes are available, and by the start of this work, many dozens are projected). As examples of Ig-like folds, fibronectin and other beta-sandwich proteins are found in algae. For each protein we will: (i) experimentally measure the stability of the proteins (single molecule force spectroscopy) and perform stability simulations, (ii) investigate how changes in bio-processing environment (surface, pH, temperature, solvent, ionic strength, additives) affect the stability and aggregation rate of the protein, (iii) measure the effect of selected mutations designed to optimise stability and aggregation propensity of the protein and (iv) apply this tool-kit to bio-therapeutic proteins to rationally modulate aggregation propensities. The proteins will be characterized in terms of 3-dimensional structure to gain insight into how they achieve temperature and acid tolerance and how these features could be more generally employed on other secreted protein targets. In particular, acid-tolerance is very poorly understood and no clear rules have yet emerged.

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