Esterases and uses thereof

Patent metadata

Field	Value
Publication number	US-10590401-B2
Application number	US-201716317178-A
Country	US
Kind code	B2
Filing date	Jul 12, 2017
Priority date	Jul 12, 2016
Publication date	Mar 17, 2020
Grant date	Mar 17, 2020

Abstract

The present invention relates to novel esterase, more particularly to esterase variants having improved activity compared to the esterase of SEQ ID NO: 1 and the uses thereof for degrading polyester containing material, such as plastic products. The esterases of the invention are particularly suited to degrade polyethylene terephthalate, and material containing polyethylene terephthalate.

First claim

The invention claimed is:

1. An esterase variant which (i) has at least 75% identity to the full length amino acid sequence set forth in SEQ ID NO: 1, and (ii) has at least one amino acid substitution at a position selected from T157, T176, S65, P58, A62, A64, L67, A68, N85, T86, R89, D91, P93, R96, G128, M131, G133, G134, T153, P154, H156, A178, P179, H183, S206, P210 or N211, wherein the positions are numbered by reference to the amino acid sequence set forth in SEQ ID NO: 1, and (iii) exhibits increased polyester degrading activity compared to an esterase of SEQ ID NO: 1.

2. The esterase variant according to claim 1 , wherein said esterase comprises at least one amino acid substitution at a position selected from T157, T176, S65 or N211, wherein the positions are numbered by reference to the amino acid sequence set forth in SEQ ID NO: 1.

3. The esterase variant according to claim 1 , wherein said esterase comprises at least one substitution selected from the group consisting of S65T, T157E/Q/N/G, T176H/N/Q and N211Q.

4. The esterase variant according to claim 1 , wherein said esterase further comprises at least one substitution at a position selected from G59, Y60, T61, D63, S66, F90, Y92, H129, G132, W155 and V177.

5. The esterase variant according to claim 4 , wherein the at least one substitution is selected from the group consisting of Y60M/F, T61M/V, D63N/Q, S66H, F90W, and Y92G/N/P/Q/T.

6. An esterase variant which (i) has at least 75% identity to the full length amino acid sequence set forth in SEQ ID NO: 1, and (ii) comprises at least one substitution or combination of substitutions selected from the group consisting of S65T, T157E/Q/N/G, T176H/N/Q, N211Q, F208W+V170I, Y92P+F208L, Y92P+F208W, T176H+F208W, V170I+A121S, V170I+A121S+S223A, F208W+T157Q, F208W+T157N, F208W+T157S, F208W+S65T and F208W+T157E, wherein the positions are numbered by reference to the amino acid sequence set forth in SEQ ID NO: 1.

7. A nucleic acid encoding an esterase variant according to claim 1 .

8. An expression cassette or vector comprising the nucleic acid of claim 7 .

9. A host cell comprising the nucleic acid of claim 7 .

10. A composition comprising the esterase variant according to claim 1 .

11. A method of producing an esterase comprising:
(a) culturing the host cell according to claim 9 under conditions suitable to express the nucleic acid encoding said esterase; and
(b) recovering said esterase from the cell culture.

12. A method of degrading a plastic product containing at least one polyester comprising (a) contacting the plastic product with an esterase according to claim 1 and thereby degrading the at least one polyester.

13. The method of claim 12 , further comprising (b) recovering monomers and/or oligomers resulting from the degradation of the at least one polyester.

14. The method of claim 12 , wherein the plastic product comprises at least one polyester selected from polyethylene terephthalate (PET), polytrimethylene terephthalate (PTT), polybutylene terephthalate (PBT), polyethylene isosorbide terephthalate (PEIT), polylactic acid (PLA), polyhydroxy alkanoate (PHA), polybutylene succinate (PBS), polybutylene succinate adipate (PBSA), polybutylene adipate terephthalate (PBAT), polyethylene furanoate (PEF), Polycaprolactone (PCL), poly(ethylene adipate) (PEA), polyethylene naphthalate (PEN) and blends/mixtures of these materials.

15. A nucleic acid encoding an esterase variant according to claim 6 .

16. An expression cassette or vector comprising the nucleic acid of claim 15 .

17. A host cell comprising the nucleic acid of claim 15 .

18. A composition comprising the esterase variant according to claim 6 .

19. A method of producing an esterase comprising:
(a) culturing the host cell according to claim 17 under conditions suitable to express the nucleic acid encoding said esterase; and
(b) recovering said esterase from the cell culture.

20. A method of degrading a plastic product containing at least one polyester comprising (a) contacting the plastic product with an esterase according to claim 6 and thereby degrading the at least one polyester.

21. The method of claim 20 , further comprising (b) recovering monomers and/or oligomers resulting from the degradation of the at least one polyester.

22. The method of claim 20 , wherein the plastic product comprises at least one polyester selected from polyethylene terephthalate (PET), polytrimethylene terephthalate (PTT), polybutylene terephthalate (PBT), polyethylene isosorbide terephthalate (PEIT), polylactic acid (PLA), polyhydroxy alkanoate (PHA), polybutylene succinate (PBS), polybutylene succinate adipate (PBSA), polybutylene adipate terephthalate (PBAT), polyethylene furanoate (PEF), Polycaprolactone (PCL), poly(ethylene adipate) (PEA), polyethylene naphthalate (PEN) and blends/mixtures of these materials.

Assignees

• Carbios

Inventors

• Tournier Vincent
• Texier Helene
• Desrousseaux Marie-Laure
• Topham Christopher
• Andre Isabelle
• Barbe Sophie
• Duquesne Sophie
• Marty Alain

Classifications

• C08J11/105

  by treatment with enzymes · CPC title

• C12Y301/01001

  Carboxylesterase (3.1.1.1) · CPC title

• C12N9/18 Primary

  Carboxylic ester hydrolases {(3.1.1)} · CPC title

• C12Y301/01074

  Cutinase (3.1.1.74) · CPC title

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title