Projects per year
Abstract
The combination of computational design and laboratory evolution is a powerful and potentially versatile strategy for the development of enzymes with new functions1,2,3,4. However, the limited functionality presented by the genetic code restricts the range of catalytic mechanisms that are accessible in designed active sites. Inspired by mechanistic strategies from small-molecule organocatalysis5, here we report the generation of a hydrolytic enzyme that uses Nδ-methylhistidine as a non-canonical catalytic nucleophile. Histidine methylation is essential for catalytic function because it prevents the formation of unreactive acyl-enzyme intermediates, which has been a long-standing challenge when using canonical nucleophiles in enzyme design6,7,8,9,10. Enzyme performance was optimized using directed evolution protocols adapted to an expanded genetic code, affording a biocatalyst capable of accelerating ester hydrolysis with greater than 9,000-fold increased efficiency over free Nδ-methylhistidine in solution. Crystallographic snapshots along the evolutionary trajectory highlight the catalytic devices that are responsible for this increase in efficiency. Nδ-methylhistidine can be considered to be a genetically encodable surrogate of the widely employed nucleophilic catalyst dimethylaminopyridine11, and its use will create opportunities to design and engineer enzymes for a wealth of valuable chemical transformations.
Original language | English |
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Pages (from-to) | 219-223 |
Journal | Nature |
Volume | 570 |
Early online date | 27 May 2019 |
DOIs | |
Publication status | Published - 27 May 2019 |
Research Beacons, Institutes and Platforms
- Manchester Institute of Biotechnology
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Dive into the research topics of 'Design and evolution of an enzyme with a non-canonical organocatalytic mechanism'. Together they form a unique fingerprint.Projects
- 1 Finished
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Manchester Synthetic Biology Research Centre for Fine and Speciality Chemicals
Scrutton, N., Azapagic, A., Balmer, A., Barran, P., Breitling, R., Delneri, D., Dixon, N., Faulon, J., Flitsch, S., Goble, C., Goodacre, R., Hay, S., Kell, D., Leys, D., Lloyd, J., Lockyer, N., Martin, P., Micklefield, J., Munro, A., Pedrosa Mendes, P., Randles, S., Salehi Yazdi, F., Shapira, P., Takano, E., Turner, N. & Winterburn, J.
14/11/14 → 13/05/20
Project: Research