Reductive Enzyme Cascades for Valorization of Polyethylene Terephthalate Deconstruction Products

Abstract

To better incentivize the collection of plastic wastes, chemical transformations must be developed that add value to plastic deconstruction products. Polyethylene terephthalate (PET) is a common plastic whose deconstruction through chemical or biological means has received much attention. However, a limited number of alternative products have been formed from PET deconstruction, and only a small share could serve as building blocks for alternative materials or therapeutics. Here, we demonstrate the production of useful monoamine and diamine building blocks from known PET deconstruction products. We achieve this by designing one-pot biocatalytic transformations that are informed by the substrate specificity of an ω-transaminase and diverse carboxylic acid reductases (CAR) toward PET deconstruction products. We first establish that an ω-transaminase from Chromobacterium violaceum (cvTA) can efficiently catalyze amine transfer to potential PET-derived aldehydes to form monoamine para-(aminomethyl)benzoic acid (pAMBA) or diamine para-xylylenediamine (pXYL). We then identified CAR orthologs that could perform the bifunctional reduction of terephthalic acid (TPA) to terephthalaldehyde or the reduction of mono-(2-hydroxyethyl) terephthalic acid (MHET) to its corresponding aldehyde. After characterizing 17 CARs in vitro, we show that the CAR from Segniliparus rotundus (srCAR) had the highest observed activity on TPA. Given these elucidated substrate specificity results, we designed modular enzyme cascades based on coupling srCAR and cvTA in one pot with enzymatic cofactor regeneration. When we supply TPA, we achieve a 69 ± 1% yield of pXYL, which is useful as a building block for polymeric materials. When we instead supply MHET and subsequently perform base-catalyzed ester hydrolysis, we achieve 70 ± 8% yield of pAMBA, which is useful for therapeutic applications and as a pharmaceutical building block. This work expands the breadth of products derived from PET deconstruction and lays the groundwork for eventual valorization of waste PET to higher-value chemicals and materials.

Genome engineering allows selective conversions of terephthalaldehyde to multiple valorized products in bacterial cells

“…Amines were derivatized with ortho ‐pthalaldehyde and 3‐mercaptopropionic acid and identified using reverse‐phase high‐performance liquid chromatography (RP‐HPLC) with an Agilent 1260 Infinity with a Zorbax Eclipse Plus‐C18 column with a guard column installed. The methods for detection of TPAL, 4HMB, 4HMBA, BDM, and pXYL used were described in Gopal et al 14 …”

“…In recent years, there have been several examples of platform chemicals produced biocatalytically from TPA, including gallic acid, pyrogallol, catechol, muconic acid, vanillic acid, and vanillin made by live cells 12,13 . We recently identified carboxylic acid reductase enzymes that can efficiently generate the versatile dialdehyde terephthalaldehyde (TPAL) from TPA 14 . Our first step was to report the use of these enzymes in vitro, in part because it is unclear if these steps would function well in live cells given the potential instability and reactivity of TPAL 15 .…”

“…Here, we envision utilizing the dialdehyde functionality of TPAL to access potential new opportunities in biosynthetic pathways, including asymmetrically functionalized products. Aldehyde‐derived biosynthetic targets have ranged broadly, 19 including diamine polymer building blocks, 14,20 hydroxylated non‐standard amino acids, 21–25 nitro alcohols, 26 and pharmaceutical mono‐amine precursors 27–29 . Despite the biosynthetic possibilities available from aldehydes, their redox instability in cellular environments due to endogenous oxidoreductase activity remains a key challenge 30 .…”

Genome engineering allows selective conversions of terephthalaldehyde to multiple valorized products in bacterial cells

“…In recent years, there have been several examples of platform chemicals produced biocatalytically from TPA, including gallic acid, pyrogallol, catechol, muconic acid, vanillic acid and vanillin made by live cells 12,13 . We recently identified carboxylic acid reductase enzymes that can efficiently generate the versatile dialdehyde terephthalaldehyde (TPAL) from TPA 14 . Our first step was to report the use of these enzymes in vitro, in part because it is unclear if these steps would function well in live cells given the potential instability and reactivity of TPAL 15 .…”

“…Here, we envision utilizing the dialdehyde functionality of TPAL to access potential new opportunities in biosynthetic pathways, including asymmetrically functionalized products. Aldehyde-derived biosynthetic targets have ranged broadly 19 , including diamine polymer building blocks 14,20 , hydroxylated non-standard amino acids [21][22][23][24][25] , nitro alcohols 26 , and pharmaceutical mono-amine precursors [27][28][29] . Despite the biosynthetic possibilities available from was not certified by peer review) is the author/funder.…”