Pterin-dependent biocatalysts and uses thereof

Patent metadata

Field	Value
Publication number	US-11085060-B2
Application number	US-201515541114-A
Country	US
Kind code	B2
Filing date	Dec 31, 2015
Priority date	Jan 2, 2015
Publication date	Aug 10, 2021
Grant date	Aug 10, 2021

Abstract

Provided herein are biocatalysts and systems thereof for pterin-dependent enzymes and pathways and methods of making and using the same. Provided herein in some embodiments are biocatalysts having a pterin source and a pterin-dependent enzymatic pathway biologically coupled to the pterin source. Tetrahydrobiopterin (referred to herein as BH4 or BH 4) can be the pterin source. The BH4 can be synthesized by a tetrahydrobiopterin synthesis pathway. The tetrahydrobiopterin synthesis pathway can include a GTP cyclohydrase; a pyruvoyl tetrahydropterin synthase; a sepiapterin reductase, and/or any combination thereof. The biocatalyst can further contain a pterin-dependent enzymatic pathway. The pterin-dependent enzymatic pathway can be amino acid mono-oxygenase, phenylalanine hydroxylase, tryptophan hydroxylase, tyrosine hydroxylase, nitric oxide synthase, alkylglycerol monooxygenase, and/or any combination thereof.

First claim

We claim:

1. A method of biocatalysis comprising the steps of:
providing a metabolite of lignin to a biocatalyst comprising a pterin-dependent enzymatic pathway in the presence of a tetrahydrobiopterin source, wherein the pterin-dependent enzymatic pathway comprises an amino acid mono-oxygenase, and
wherein the metabolite of lignin is one of:

and
wherein the amino acid mono-oxygenase is encoded by a nucleotide sequence comprising SEQ ID NO: 14 or SEQ ID NO: 15.

2. The method of claim 1 , wherein the biocatalyst further comprises a tetrahydrobiopterin recycling pathway, the tetrahydrobiopterin recycling pathway comprising:
a pterin-4a-carbinolamine dehydratase; and
a dihydropterin reductase.

3. The method of claim 1 , wherein the metabolite of lignin is:

4. The method of claim 1 , wherein the tetrahydrobiopterin source is a tetrahydrobiopterin synthesis pathway comprising:
a GTP cyclohydrolase;
a pyruvoyl tetrahydropterin synthase; and
a sepiapterin reductase.

5. The method of claim 1 , wherein the biocatalyst further comprises an aromatic-l-amino acid decarboxylase.

6. The method of claim 1 , wherein the biocatalyst further comprises a terpene alkaloid synthase.

7. A method of biocatalysis comprising the steps of:
providing a metabolite of lignin to a biocatalyst comprising a pterin-dependent enzymatic pathway in the presence of a tetrahydrobiopterin source, wherein the pterin-dependent enzymatic pathway comprises an amino acid mono-oxygenase, and
wherein the metabolite of lignin is:

8. The method of claim 2 , wherein the pterin-4a-carbinolamine dehydratase is encoded by a nucleotide sequence comprising SEQ ID NO: 12; and
the dihydropterin reductase is encoded by a nucleotide sequence comprising SEQ ID NO: 13.

9. The method of claim 4 , wherein the GTP cyclohydrolase is encoded by a nucleotide sequence comprising SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4;
the pyruvoyl tetrahydropterin synthase is encoded by a nucleotide sequence comprising SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, or SEQ ID NO: 8; and
the sepiapterin reductase is encoded by a nucleotide sequence comprising SEQ ID NO: 9, SEQ ID NO: 10, or SEQ ID NO: 11.

10. The method of claim 5 , wherein the aromatic-l-amino acid decarboxylase is encoded by a nucleotide sequence comprising SEQ ID NO: 16.

11. The method of claim 6 , wherein the terpene alkaloid synthase is encoded by a nucleotide sequence comprising SEQ ID NO: 17.

Assignees

• Georgia Tech Res Inst

Inventors

• Peralta-Yahya Pamela
• Ehrenworth Amy M

Classifications

• C12Y402/01096

  4a-Hydroxytetrahydrobiopterin dehydratase (4.2.1.96) · CPC title

• C12Y305/04016

  GTP cyclohydrolase I (3.5.4.16) · CPC title

• C12N9/0036

  acting on NADH or NADPH (1.6) · CPC title

• C12Y101/01153

  Sepiapterin reductase (L-erythro-7,8-dihydrobiopterin forming) (1.1.1.153) · CPC title

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title