Engineered microorganisms for the deconstruction of polymers

Patent metadata

Field	Value
Publication number	US-12371718-B2
Application number	US-202117198230-A
Country	US
Kind code	B2
Filing date	Mar 10, 2021
Priority date	May 15, 2018
Publication date	Jul 29, 2025
Grant date	Jul 29, 2025

Abstract

Disclosed herein are methods and compositions for catalytic glycolysis to deconstruct PET to bis(2-hydroxyethyl) terephthalate (BHET). For BHET conversion to terephthalate and ethylene glycol, we engineer Pseudomonas putida KT2440 with PETase and MHETase enzymes from Ideonella sakaiensis . We further engineer P. putida to convert terephthalate to a performance-advantaged bioproduct, β-ketoadipic acid, and for improved utilization of ethylene glycol, a byproduct of BHET catabolism. In a bioreactor, we produce 15.1±0.6 g/L of β-ketoadipic acid (βKA) from BHET at 76±3% molar yield. Lastly, we demonstrate conversion of catalytically depolymerized PET to βKA. Overall, this work highlights the potential of tandem catalytic deconstruction and biological conversion as a means to upcycle waste PET.

First claim

What is claimed is:

1. A genetically modified Pseudomonas sp. organism, comprising:
a nucleic acid sequence encoding a functional Ideonella sakaiensis PETase comprising a nucleic acid sequence encoding for a first secretion signal peptide; and
a nucleic acid sequence encoding a functional Ideonella sakaiensis MHETase comprising a nucleic acid sequence encoding for a second secretion signal peptide;
wherein the genetically modified organism metabolizes bis(2-hydroxyethyl) terephthalate (BHET) to produce BHET deconstruction products.

2. The genetically modified organism of claim 1 , wherein both nucleic acid sequences are codon optimized.

3. The genetically modified organism of claim 1 , wherein both nucleic acid sequences are incorporated into the genome of the genetically modified organism.

4. The genetically modified organism of claim 1 , wherein the nucleic acid sequence encoding a first secretion signal peptide encodes for a signal peptide with greater than 90% sequence identity to SEQ ID NO: 17; and wherein the nucleic acid sequence encoding for a second secretion signal peptide encodes for a signal peptide with greater than 90% sequence identity to SEQ ID NO: 19.

5. The genetically modified organism of claim 1 , wherein the organism is Pseudomonas putida.

6. The genetically modified organism of claim 1 , wherein the BHET deconstruction products comprise at least one of mono-(2-hydroxyethyl) terephthalate, terephthalate, ethylene glycol, ß-ketoadipate, or muconate.

7. A method comprising contacting poly (ethylene terephthalate) (PET) with the genetically modified organism of claim 1 to produce PET deconstruction products.

8. The method of claim 7 , wherein the contacting is performed in minimal salt medium.

Assignees

• Alliance Sustainable Energy
• Ut Battelle Llc

Inventors

• Beckham Gregg Tyler
• Jayakody Thelhawadigedara Lahiru Niroshan
• Guss Adam Michael
• Mand Thomas David
• Johnson Christopher W
• Pardo Mendoza Isabel
• Zimont Werner Allison Jean

Classifications

• C12N9/18 Primary

  Carboxylic ester hydrolases {(3.1.1)} · CPC title

• C12Y301/01

  Carboxylic ester hydrolases (3.1.1) · CPC title

• C12P7/18

  polyhydric · CPC title

• C12P7/44

  Polycarboxylic acids · CPC title

• Y02P20/582

  Recycling of unreacted starting or intermediate materials · CPC title