Computational redesign of enzymes for regio- and enantioselective hydroamination

Abstract

Introduction of innovative biocatalytic processes offers great promise for applications in green chemistry. However, owing to limited catalytic performance, the enzymes harvested from nature's biodiversity often need to be improved for their desired functions by time-consuming iterative rounds of laboratory evolution. Here we describe the use of structure-based computational enzyme design to convert Bacillus sp. YM55-1 aspartase, an enzyme with a very narrow substrate scope, to a set of complementary hydroamination biocatalysts. The redesigned enzymes catalyze asymmetric addition of ammonia to substituted acrylates, affording enantiopure aliphatic, polar and aromatic β-amino acids that are valuable building blocks for the synthesis of pharmaceuticals and bioactive compounds. Without a requirement for further optimization by laboratory evolution, the redesigned enzymes exhibit substrate tolerance up to a concentration of 300 g/L, conversion up to 99%, β-regioselectivity >99% and product enantiomeric excess >99%. The results highlight the use of computational design to rapidly adapt an enzyme to industrially viable reactions.

Computational redesign of a PETase for plastic biodegradation by the GRAPE strategy

“…Ameliorated biodegradation of other plastics, such as poly(butylene terephthalate) (PBT) and poly(ethylene 2,6-naphthalenedicarboxylate) (PEN), suggested DuraPETase a broad ability to degrade semiaromatic polyesters. To evaluate 45 the underlying molecular mechanism for enhanced performance, a three-dimensional crystal structure of the variant was determined; this structure highlighted a fine-tuned 'aromatic tunnel' flanked by synergistic hydrophobic interactions.…”

Efficient production of phenylpropionic acids by an amino‐group‐transformation biocatalytic cascade

“…However, due to the low activity of Tc PAM C107S, L104A , ( R )‐β‐tyrosine derivatives were produced in very low concentration. To date, the maximum titer of ( R )‐β‐amino acids is 355.7 g/L of ( R )‐β‐aminobutanoic acid, which was produced by the asymmetric addition of ammonia to crotonic acid catalyzed by a computational redesigned enzyme AspB B19 (Li et al, ). Thus, to increase titer of ( R )‐β‐tyrosine derivatives, more research is needed to increase the stability and activity of tyrosine AMs.…”

Broadening the Scope of Biocatalysis in Sustainable Organic Synthesis

“…[59] Nowadays, protein engineering strategies for improving enzyme performance generally involve combining computeraided design with directed evolution. Interestingly,J anssen and co-workers [60] recently reported the computational redesign of aspartase,w hich catalyzes the reversible deamination of a-aspartate to fumarate and ammonia, affordinganeffective catalystf or the enantioselective b-hydroamination of a,bunsaturated carboxylic acids (Scheme 5). Aspartase is ah ighly selectivee nzyme from primary metabolism with no known promiscuous activity (see Section 6) making it anotoriouslyd ifficult candidate for directed evolution, requiring the screening of large numbers of variants.…”