Microorganisms for the production of 5-hydroxytryptophan
OwnerEric Michael Knight, Jiangfeng Zhu +2inventors
InventorsEric Michael Knight, Jiangfeng Zhu, Jochen Förster, Hao Luo
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Abstract
Recombinant microbial cells and methods for producing 5-hydroxytryptophan (5HTP) using such cells are described. More specifically, the recombinant microbial cell comprises an exogenous gene encoding an L-tryptophan hydroxylase, and means for providing tetrahydrobiopterin (THB). Related sequences and vectors for use in preparing such recombinant microbial cells are also described.
Claims18
A recombinant microbial cell comprising an exogenous nucleic acid sequence encoding an L-tryptophan hydroxylase (TPH) (EC 1.14.16.4), and exogenous nucleic acid sequences encoding enzymes of at least one pathway for producing tetrahydrobiopterin (THB).
The recombinant microbial cell of any one of the preceding claims, comprising exogenous nucleic acid sequences encoding
- (a) optionally, a GTP cyclohydrolase I (EC 3.5.4.16);
- (b) a 6-pyruvoyl-tetrahydropterin synthase (EC 4.2.3.12);
- and
- (c) a sepiapterin reductase (EC 1.1.1.153).
The recombinant microbial cell of any one of the preceding claims, comprising exogenous nucleic acid sequences encoding
- (a) a 4a-hydroxytetrahydrobiopterin dehydratase (EC 4.2.1.96);
- and
- (b) optionally, a dihydropteridine reductase (EC 1.5.1.34).
The recombinant microbial cell of any one of the preceding claims, wherein each one of said exogenous nucleic acid sequences is operably linked to an inducible, a regulated or a constitutive promoter.
The recombinant microbial cell of any one of the preceding claims, which comprises a mutation providing for reduced tryptophanase activity.
The recombinant microbial cell of any one of the preceding claims, which is derived from a microbial host cell which is a bacterial cell, a yeast host cell, a filamentous fungal cell, or an algeal cell.
The recombinant microbial cell of any one of the preceding claims, which is an Escherichia coli cell.
The recombinant microbial cell of any one of the preceding claims, wherein the L-tryptophan hydroxylase comprises the amino acid sequence of SEQ ID NO:9.
A vector comprising nucleic acids encoding an L-tryptophan hydroxylase, a 4a-hydroxytetrahydrobiopterin dehydratase, and a dihydropteridine reductase.
A method of producing a recombinant microbial cell, comprising transforming a microbial host cell with one or more vectors comprising nucleic acid sequences encoding
- (a) an L-tryptophan hydroxylase (EC 1.14.16.4);
- (b) a GTP cyclohydrolase I (EC 3.5.4.16);
- (c) a 6-pyruvoyl-tetrahydropterin synthase (EC 4.2.3.12);
- (d) a sepiapterin reductase (EC 1.1.1.153);
- (e) a 4a-hydroxytetrahydrobiopterin dehydratase (EC 4.2.1.96);
- and
- (f) a dihydropteridine reductase (EC 1.5.1.34),
- each one of said nucleic acid sequences being operably linked to an inducible, a regulated or a constitutive promoter, thereby obtaining the recombinant microbial cell.
Dependencies are inferred from the claim text.