Cell factory having improved iron-sulfur cluster delivery
AssigneeBiosyntia Aps
InventorsHans Jasper Genee, Anne Pihl Bali, Nils Myling-Petersen
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Abstract
The invention provides a genetically modified bacterial cell capable of improved iron-sulfur cluster delivery, characterized by a modified gene encoding a mutant Iron Sulfur Cluster Regulator (IscR) as well as one or more transgenes encoding polypeptides that enhance the biosynthesis of either biotin, lipoic acid or thiamine. The invention provides a method for producing either biotin, lipoic acid or thiamine using the genetically modified bacterium of the invention; as well as for the use of the genetically modified bacterial cell for either biotin, lipoic acid or thiamine production.
Claims18
A genetically modified bacterium wherein the bacterium has an enhanced production of biotin or lipoic acid or thiamine;
- wherein said bacterium comprises:
- a. a genetically modified endogenous iscR gene encoding a mutant
- IscR polypeptide, wherein the amino acid sequence of said mutant IscR polypeptide has at least 80% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID No:
- 2, 4, 6, 8, 10, 12 and 14, and wherein said amino acid sequence has at least one amino acid substitution selected from the group consisting of:
- i) L15X, C92X, C98X, C104X, and H 107X;
- wherein X is any amino acid other than the corresponding amino acid residue in SEQ ID No.:
- 2, 4, 6, 8, 10, 12 and 14;
- and
- b. at least one transgene encoding a polypeptide selected from the group consisting of:
- ii) a polypeptide having biotin synthase activity (EC 2.8.1.6),
- iii) a polypeptide having lipoic acid synthase activity (EC 2.8.1.8),
- iv). a polypeptide having HMP-P synthase activity (EC 4.1.99.17), and
- v). a polypeptide having tyrosine lyase activity (EC 4.1.99.19).
The method of a genetically modified gene encoding a mutant iscR polypeptide to enhance production of any one of biotin, lipoic acid or thiamine in a genetically modified bacterium, wherein said bacterium comprises and expresses at least one transgene encoding a polypeptide selected from the group consisting of:
- i) a polypeptide having biotin synthase activity (EC 2.8.1.6),
- ii) a polypeptide having lipoic acid synthase activity (EC 2.8.1.8),
- iii) a polypeptide having HMP-P synthase activity (EC 4.1.99.17), and
- iv) a polypeptide having tyrosine lyase activity (EC 4.1.99.19), and
- wherein said genetically modified gene is an endogenous iscR gene encoding a mutant IscR polypeptide, wherein the amino acid sequence of said mutant IscR polypeptide has at least 80% amino acid sequence identity to SEQ ID No.:
- 2, 4, 6, 8, 10, 12 and 14, and wherein the amino acid sequence has at least one amino acid substitution selected from the group consisting of:
- L15X, C92X, C98X, C104X, and H 107X;
- wherein X is any amino acid other than the corresponding amino acid residue in SEQ ID No 2, 4, 6, 8, 10, 12 and 14.
Dependencies are inferred from the claim text.