Abstract
Although C–N bonds are ubiquitous in natural products, pharmaceuticals and agrochemicals, biocatalysts forging these bonds with high atom-efficiency and enantioselectivity have been limited to a few select enzymes. In particular, ammonia lyases have emerged as powerful catalysts to access C–N bond formation via hydroamination. However, the use of ammonia lyases is rather restricted due to their narrow synthetic scope. Herein, we report the computational redesign of aspartase, a highly specific ammonia lyase, to yield C–N lyases with cross-compatibility of non-native nucleophiles and electrophiles. A wide range of non-canonical amino acids (ncAAs) are afforded with excellent conversion (up to 99%), regioselectivity >99% and enantioselectivity >99%. The process is scalable under industrially relevant protocols (exemplified in kilogram-scale synthesis) and can be facilely integrated in cascade reactions (demonstrated in the synthesis of β-lactams with N-1 and C-4 substitutions). This versatile and efficient C–N lyase platform supports the preparation of ncAAs and their derivatives, and will present opportunities in synthetic biology. Ammonia lyases are powerful catalysts to access C–N bond formation via hydroamination, but show a narrow synthetic scope. Now, by computational redesign of an aspartase, a C–N lyase is developed that shows cross-compatibility of non-native nucleophiles and electrophiles expanding the synthetic scope.
Highlights
Outside of the twenty proteinogenic amino acids that serve as the foundational building blocks of life, there are manifold noncanonical amino acids that exhibit diverse physiological functions and are extensively used as intermediates for bioactive products[1]
The physical and chemical properties of synthetic peptides and proteins can be selectively manipulated through the incorporation of noncanonical amino acids (ncAAs), which has contributed to understanding biological macromolecules[3] and the development of novel therapeutics[4] and high-performing biocatalysts[5,6]
We envisioned that the aspartase from Bacillus sp. YM55-1 (AspB) variants with a thoroughly reshaped active site could achieve cross addition of a large variety of nucleophilic amines to unsaturated acids, providing a C-N lyase platform that possesses unique and attractive properties for the biocatalytic preparation of a myriad of practically relevant ncAAs and their demand-tailored derivatives
Summary
Outside of the twenty proteinogenic amino acids that serve as the foundational building blocks of life, there are manifold noncanonical amino acids (ncAAs) that exhibit diverse physiological functions and are extensively used as intermediates for bioactive products[1]. YM55-1 (AspB)[20], we attempted to tackle the longstanding challenge of creating C-N lyases with crosscompatibility of nonnative substrates by performing computational protocols that allow large jumps in function to traverse inactive sequence space along the fitness landscape.
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