We report for the first time the combination of directed evolution focused on enhancing and reversing the stereoselectivity of an enzyme with Cu(I)-mediated click chemistry(CuAAC),providing an asymmetric click approac...We report for the first time the combination of directed evolution focused on enhancing and reversing the stereoselectivity of an enzyme with Cu(I)-mediated click chemistry(CuAAC),providing an asymmetric click approach for versatile chiral triazoles products.In this study,the halohydrin dehalogenase HheG was used as the enzyme which was evolved to induce a stereoselective ring-opening reaction of cyclic epoxides in the presence of NaN3 with the formation of chiral azido products.Two mutants of opposite stereopreference were generated,which convert cyclohexene oxide as well as cycloheptene oxide to(1S,2S)-2-azidocyclohexanol,(1R,2R)-2-azidocyclohexanol,(1S,2S)-2-azidocycloheptanol and(1R,2R)-2-azidocycloheptanol with essentially high stereoselectivity.The chiral products were then subjected to CuAAC in reactions with structurally different alkynes.Since HheG was found to be compatible with Cu(I),the process was also performed successfully in a unique 2-step one-pot process leading to various chiral triazoles.In order to understand the enhancement and reversal of the evolved enantioselectivity,QM and MD computations were performed.This approach harnesses the modifiability and high stereoselectivity of the evolved biocatalysts in combination with click chemistry.It holds great potential for diverse fields,particularly in the area of pharmaceuticals.展开更多
基金support from the National Key Research and Development Program of China(No.2023YFA0914100/2023YFA0914102)the National Natural Science Foundation of China(Nos.22077029 and 22034002)+2 种基金the Science Fund for Distinguished Young Scholars of Hunan Province(No.2021JJ10034)support from the National Natural Science Foundation of China(No.22276049)M.M.acknowledge the support from the National Natural Science Foundation of China(No.22276050).
文摘We report for the first time the combination of directed evolution focused on enhancing and reversing the stereoselectivity of an enzyme with Cu(I)-mediated click chemistry(CuAAC),providing an asymmetric click approach for versatile chiral triazoles products.In this study,the halohydrin dehalogenase HheG was used as the enzyme which was evolved to induce a stereoselective ring-opening reaction of cyclic epoxides in the presence of NaN3 with the formation of chiral azido products.Two mutants of opposite stereopreference were generated,which convert cyclohexene oxide as well as cycloheptene oxide to(1S,2S)-2-azidocyclohexanol,(1R,2R)-2-azidocyclohexanol,(1S,2S)-2-azidocycloheptanol and(1R,2R)-2-azidocycloheptanol with essentially high stereoselectivity.The chiral products were then subjected to CuAAC in reactions with structurally different alkynes.Since HheG was found to be compatible with Cu(I),the process was also performed successfully in a unique 2-step one-pot process leading to various chiral triazoles.In order to understand the enhancement and reversal of the evolved enantioselectivity,QM and MD computations were performed.This approach harnesses the modifiability and high stereoselectivity of the evolved biocatalysts in combination with click chemistry.It holds great potential for diverse fields,particularly in the area of pharmaceuticals.
