Optimized microbial cells for production of melatonin and other compounds
AssigneeDanmarks Tekniske Universitet
InventorsHao Luo, Jochen Förster
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Abstract
Described herein are recombinant microbial host cells comprising biosynthetic pathways and their use in producing oxidation products and downstream products, e.g., melatonin and related compounds, as well as enzyme variants, nucleic acids, vectors and methods useful for preparing and using such cells. In specific aspects, the present invention relates to monooxygenases, e.g., amino acid hydroxylases, with a modified cofactor-dependency, and to enzyme variants and microbial cells providing for an improved supply of cofactors.
Claims20
A variant of a tryptophan hydroxylase (TPH), the variant comprising
- (a) a segment corresponding to residues E147 to T460 of Homo sapiens TPH having the sequence of SEQ ID NO:3,
- (b) an N-terminal methionine residue, and
- (c) a mutation in at least one residue corresponding to residues N242 and P244 in SEQ ID NO:3, wherein the mutation in the residue corresponding to residue N242 is the amino acid substitution 242I and the mutation in the residue corresponding to P244 is selected from 244C, 244D, 244L and 244Q,
- wherein the variant is a variant of a Homo sapiens TPH having the sequence of SEQ ID NO:3, and wherein the segment in the variant has at least about 80% sequence identity to the segment in the native sequence, and
- wherein the variant provides for a tryptophan hydroxylation activity which is at least 110% of that of Homo sapiens TPH having the sequence of SEQ ID NO:3.
A TPH having the sequence of SEQ ID NO 13, further comprising E2K, N97I and P99C mutations.
Dependencies are inferred from the claim text.