Repurposing Carboxylic Acid Reductase for Amide Bond Formation

Abstract

As about one-quarter of marketed drugs contain at least one amide bond, amine acylation is the most commonly performed reaction in the pharmaceutical industry. There is therefore a high demand for sustainable alternatives to classical amidation methods. Carboxylic acid reductase (CAR) is a multidomain enzyme, catalysing the reduction of carboxylic acids to aldehydes via a multistep process utilizing the cofactors ATP and NADPH. It has previously been discovered that the acyl adenylate intermediate can be intercepted in this process by adding an excess of amine nucleophile, forming an amide bond. Here, we have engineered the CAR enzyme specifically for amidation (CAR-A), improving conversions for amide bond formation. Furthermore, a polyphosphate kinase (CHU) was implemented to recycle the ATP cofactor from cheap polyphosphate. This system was then exploited for the mono acylation of symmetrical diamines. Mono-acylated diamines are common motifs in numerous pharmaceuticals and traditional synthesis methods rely on wasteful protection/deprotection strategies. CAR-A is able to selectively mono-acylate diamines without any diacylated product being detected. Subsequently, we were interested in using this system for the synthesis of N-alkanoyl-N-methylglucamides (MEGAs). MEGAs are non-ionic surfactants widely used in biochemical and pharmaceutical applications because of their non-toxic properties. By selectively coupling fatty acids with amino-alcohols this method improves on previously reported methods as it shows no competing esterification reaction products and is carried out under aqueous conditions. This work establishes a biocatalytic amidation method that is able to couple a wide range of carboxylic acids and amines under mild reaction conditions.

Date of Award	1 Aug 2023
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Sabine Flitsch (Supervisor) & Nicholas Turner (Supervisor)