Data-efficient distal engineering of fluorinase using zero-shot models
Harding-Larsen, D.; Lax, B. M.; Weingarten, C. K.; Sako, A.; Mazurenko, S.; Welner, D. H.
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Fluorinases have high potential for industrial biofluorination but any applications have been precluded by low catalytic efficiency and resistance to active site engineering. In this work, we employed PRIZM, a computational workflow utilizing an existing low-N dataset and zero-shot models for in silico prediction of activity-enhancing mutations at distal sites. The combination of these predictions with expert opinion led to the identification of 21 fluorinase mutants with enhanced relative activities, while 3 variants showed increased melting temperatures. A mutation in the hexameric interface, K237R, resulted in the largest stability gain, a more than 3.2-fold improvement in catalytic efficiency at 57{degrees}C, and an 8-fold increase in relative activity at 62{degrees}C. These results highlight the potential of distal fluorinase engineering for improving properties required to realize its industrial applications.
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