Thioester-mediated biocatalytic amide bond synthesis with in situ thiol recycling

The activation of carboxylic acids to thioesters plays an important role in biology. However, biochemical studies and biotechnological applications are hampered by a general lack of access to thioesters, especially those based on Coenzyme A (CoA-SH). Here we show a generic thioester recycling enzyme by exploiting the promiscuous activity of a carboxylic acid reductase (CARsr). The adenylation domain of CARsr (CARsr-A) catalyses the conversion of a wide range of carboxylic acids to acyl-S-Coenzyme A and other thioesters in good yields. CARsr-A was used in situ as part of a recycling system to regenerate thioesters for acyl-S-Coenzyme A-dependent enzymes in one-pot reactions. This concept of thioester recycling is demonstrated with a range of acyltransferases that allow the formation of diverse amides and the non-native acylation of lysine side chains in a histone-derived peptide using the epigenetic writer, lysine acetyltransferase HATp300. Overall, these results establish a generic platform for thioester formation towards amide formation and beyond.

Automated summary

The activation of carboxylic acids to thioesters, especially Coenzyme A-based ones, is important but challenging. A generic thioester recycling enzyme, CARsr-A, was developed to catalyze the conversion of carboxylic acids to thioesters. This recycling system can regenerate thioesters for various acyl-S-Coenzyme A-dependent enzymes and enable the formation of diverse amides and non-native acylation reactions.