(250c) Reductive Enzyme Cascades for Valorization of PET Deconstruction Products Guided By the Specificity of Carboxylic Acid Reductases

To better incentivize the chemical recycling of plastic wastes, new chemical transformations must be developed. Polyethylene terephthalate (PET) is a recalcitrant plastic whose deconstruction through chemical and biological means has recently received much attention. However, a limited number of functionalized products have been formed from the PET monomer terephthalic acid (TPA). In this work, we present the design of a biocatalytic cascade to convert TPA, a dicarboxylate, to its corresponding diamine, para-xylylenediamine (pXYL), by constructing a multienzyme cascade in a cell-free environment. Design of our cascade first begins by probing the substrate specificity of natural carboxylic acid reductases (CARs). CARs catalyze the selective 2-electron reduction of carboxylic acids to aldehydes. In addition to screening many existing and novel CARs towards a range of carboxylate-containing PET deconstruction products, we also compared rates of reduction to the associated aldehydes in a system which included enzymatic NADPH and ATP regeneration. While our highest performing CAR was not able to completely convert 5mM of TPA to terephthalaldehyde (TPAL) at our chosen endpoint of 24-hours, coupling CAR to an ω-transaminase, which converts aldehydes to amines in the presence of an amine donor, drove the conversion of TPA to near completion, and resulted in a high yield and selectivity of the target diamine, pXYL. By combining chemical depolymerization with enzymatic functionalization, we show, to our knowledge, the first reports of CAR specificity towards TPA and biocatalytic synthesis of TPAL and pXYL. This work lays the foundation for eventual valorization of waste PET to higher value materials, such as nonisocyanate polyurethanes, that can be made from pXYL.