Design and Evolution of Photo-Enzymes for Stereoselective Transformations of Nitrogen Radicals
Lead Research Organisation:
University of Manchester
Department Name: Chemistry
Abstract
Enzymes are exceptionally powerful catalysts that recognize molecular substrates and process them in active sites. They are generally built from just 20 amino acids, and their catalytic machinery is typically assembled from chemical groups in the amino-acid side chains. But fewer than half of these side chains contain functional groups that can participate in enzyme catalytic cycles, which severely restricts the range of mechanisms conceivable within enzyme active sites. This raises the intriguing question of whether the catalytic repertoire of enzymes could be expanded by using an extended 'alphabet' of amino acids that offers a wider range of side chains for catalysis. In recent years our group have begun to take major strides towards achieving this ambitious vision (e.g. Nature 2019, 570, 219, J. Am. Chem. Soc. 2018, 140, 1535, J. Am. Chem. Soc. 2016, 138, 11344).
Our approach exploits engineered cellular translation components to selectively install non-canonical amino acids containing functional side chains. Genetically encoding the non-canonical functionality offers enormous advantages over alternative methods for chemically modifying protein structure: it greatly facilitates the production of well-defined, homogeneous proteins; it allows the non-canonical amino acid to be introduced at any site, in any protein scaffold; and, perhaps most significantly, it allows for rapid optimization of enzyme properties using directed evolution. Inspired by mechanistic strategies from small molecule organocatalysis, we have recently employed a combination of genetic code expansion, computational enzyme design and laboratory evolution to create enzymes that exploit non-canonical amino acids as catalytic nucleophiles (Nature 2019, 570, 219). This study now opens up new and exciting opportunities to enzyme designers and engineers which will be fully explored within this PhD studentship. Free from the constraints of the genetic code, the student will employ our advanced enzyme engineering techniques to create enzymes with functions not observed in Nature, that were previously thought inaccessible to the field of biocatalysis.
The project will specifically aim to create enzymes that promote enantioselective photo-redox transformations. In recent years, the Leonori group (project partner) have developed a wealth of valuable photo-redox processes involving the intermediate generation of nitrogen radicals. At present, these transformations produce racemic products, and the development of enantioselective versions of these reactions remains a key unresolved challenge. To address the objective, we will exploit engineered cellular translation components available in our laboratory to embed organic cofactors with suitable properties for mediating photo-induced electron transfers into designed active sites. Here we can take advantage of molecular recognition elements provided by the protein scaffold to achieve enantioselective conversions. Significantly, promising starting designs can be substantially improved through iterative rounds of directed evolution to afford highly efficient and selective photo-redox enzymes for the production of high value molecules.
Our approach exploits engineered cellular translation components to selectively install non-canonical amino acids containing functional side chains. Genetically encoding the non-canonical functionality offers enormous advantages over alternative methods for chemically modifying protein structure: it greatly facilitates the production of well-defined, homogeneous proteins; it allows the non-canonical amino acid to be introduced at any site, in any protein scaffold; and, perhaps most significantly, it allows for rapid optimization of enzyme properties using directed evolution. Inspired by mechanistic strategies from small molecule organocatalysis, we have recently employed a combination of genetic code expansion, computational enzyme design and laboratory evolution to create enzymes that exploit non-canonical amino acids as catalytic nucleophiles (Nature 2019, 570, 219). This study now opens up new and exciting opportunities to enzyme designers and engineers which will be fully explored within this PhD studentship. Free from the constraints of the genetic code, the student will employ our advanced enzyme engineering techniques to create enzymes with functions not observed in Nature, that were previously thought inaccessible to the field of biocatalysis.
The project will specifically aim to create enzymes that promote enantioselective photo-redox transformations. In recent years, the Leonori group (project partner) have developed a wealth of valuable photo-redox processes involving the intermediate generation of nitrogen radicals. At present, these transformations produce racemic products, and the development of enantioselective versions of these reactions remains a key unresolved challenge. To address the objective, we will exploit engineered cellular translation components available in our laboratory to embed organic cofactors with suitable properties for mediating photo-induced electron transfers into designed active sites. Here we can take advantage of molecular recognition elements provided by the protein scaffold to achieve enantioselective conversions. Significantly, promising starting designs can be substantially improved through iterative rounds of directed evolution to afford highly efficient and selective photo-redox enzymes for the production of high value molecules.
Planned Impact
iCAT will work with industry partners to create an holistic approach to the training of students in biocatalysis, chemocatalysis, and their process integration. Traditional graduate training typically focuses on one aspect of catalysis and this approach can severely restrict innovation and impact. Advances in technology and fundamental reaction discovery are rendering this silo-approach obsolete, and a new training modality is needed to produce the next generation of chemists and engineers who can operate across a far broader chemical continuum. iCAT will meet this challenge with a state-of-the-art CDT, equipping the next generation of scientists and engineers with the skills needed to develop future catalytic processes and create the functional molecules of tomorrow.
The UK has one of the world's top-performing chemical industries, achieving outstanding levels of growth, exports, productivity and international investment. The UK's chemical industry is a significant provider of jobs and creator of wealth, with a turnover in excess of £50 billion and a contribution of over £15 Billion of value to the UK economy [2015 figures]. iCAT will deliver highly skilled people to lead this industry across its various sectors, achieving impact through the following actions:
1. Equip the next generation of science and engineering leaders with the interdisciplinary skills and knowledge needed to work across the bio and chemo catalytic remit and build the functional molecules we need to structure society.
2. Provide a highly skilled workforce and research base, skilled in the latest methodologies, strategies and techniques of catalysis and engineering that is crucial for the UK's Chemical Industry.
3. Build the critical mass necessary to support effective cohort-based training in a world-class research environment.
4. Develop and disseminate new catalytic technologies and processes that will be taken up by industrial and academic teams around the world.
5. Encourage Industry to promote research challenges within the CDT that are of core relevance to their business.
6. Provide cohesion in the integration of biocatalysis, engineering and chemocatalysis to create a more unified voice for strategic dialogue with industry, funders and policy makers, and more generally outreach and public engagement.
7. Draw-in and bring together Industrial partners to facilitate future Industrial collaborations.
8. Benefit Industrial scientists through interactions with the CDT (e.g. training and supervisory experience, exposure to cutting-edge synthesis and catalysis etc).
9. Link with other activities in the landscape: bringing unique expertise in catalysis to, for example, externally-funded University-led initiatives, EPRSC Grand Challenge Networks, and the National Catalysis Hub.
The UK has one of the world's top-performing chemical industries, achieving outstanding levels of growth, exports, productivity and international investment. The UK's chemical industry is a significant provider of jobs and creator of wealth, with a turnover in excess of £50 billion and a contribution of over £15 Billion of value to the UK economy [2015 figures]. iCAT will deliver highly skilled people to lead this industry across its various sectors, achieving impact through the following actions:
1. Equip the next generation of science and engineering leaders with the interdisciplinary skills and knowledge needed to work across the bio and chemo catalytic remit and build the functional molecules we need to structure society.
2. Provide a highly skilled workforce and research base, skilled in the latest methodologies, strategies and techniques of catalysis and engineering that is crucial for the UK's Chemical Industry.
3. Build the critical mass necessary to support effective cohort-based training in a world-class research environment.
4. Develop and disseminate new catalytic technologies and processes that will be taken up by industrial and academic teams around the world.
5. Encourage Industry to promote research challenges within the CDT that are of core relevance to their business.
6. Provide cohesion in the integration of biocatalysis, engineering and chemocatalysis to create a more unified voice for strategic dialogue with industry, funders and policy makers, and more generally outreach and public engagement.
7. Draw-in and bring together Industrial partners to facilitate future Industrial collaborations.
8. Benefit Industrial scientists through interactions with the CDT (e.g. training and supervisory experience, exposure to cutting-edge synthesis and catalysis etc).
9. Link with other activities in the landscape: bringing unique expertise in catalysis to, for example, externally-funded University-led initiatives, EPRSC Grand Challenge Networks, and the National Catalysis Hub.
Organisations
People |
ORCID iD |
Anthony Green (Primary Supervisor) | |
Jonathan Trimble (Student) |
Studentship Projects
Project Reference | Relationship | Related To | Start | End | Student Name |
---|---|---|---|---|---|
EP/S023755/1 | 31/03/2019 | 29/09/2027 | |||
2279462 | Studentship | EP/S023755/1 | 30/09/2019 | 29/09/2023 | Jonathan Trimble |