Recombinant bacteria engineered to treat diseases associated with methionine metabolism and methods of use thereof
OwnerJian-Rong Gao, Jillian Marie Meansinventors
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Abstract
The present disclosure provides host cells, e.g., bacterial cells, that comprise a methionine decarboxylase enzyme for the treatment of diseases and disorders associated with methionine metabolism in a subject. The disclosure further provides pharmaceutical compositions and methods of treating disorders associated with methionine metabolism, such as homocystinuria.
Claims82
A recombinant bacterial cell comprising a heterologous methionine decarboxylase (metDC) gene operably linked to a promoter, wherein the heterologous metDC gene encodes a polypeptide that has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1136, 1137, or 1138.
A recombinant bacterial cell comprising a heterologous methionine decarboxylase (metDC) gene operably linked to a promoter, wherein the heterologous metDC gene comprises a sequence that has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1133, 1134, or 1135.
The recombinant bacterial cell of any one of the previous claims, further comprising genetic modification that reduces export of methionine from the bacterial cell.
The recombinant bacterial cell of any one of the previous claims, further comprising an insertion, deletion or mutation of an endogenous phage gene.
The recombinant bacterial cell of any one of the previous claims, further comprising a modified endogenous colibactin island.
The recombinant bacterial cell of any one of the previous claims, further comprising an auxotrophic modification.
The recombinant bacterial cell of any one of the previous claims, wherein the recombinant bacterial cell comprises two or three copies of the heterologous metDC gene.
The recombinant bacterial cell of any one of the previous claims, wherein the promoter is an inducible promoter.
The recombinant bacterial cell of any one of the previous claims, wherein the heterologous gene is located on a plasmid or a chromosome in the bacterial cell.
A recombinant bacterial cell comprising:
- a heterologous methionine decarboxylase (metDC) gene operably linked to a promoter, wherein the heterologous metDC gene encodes a polypeptide that has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1136, 1137, or 1138,
- a heterologous gene encoding a methionine importer, wherein the heterologous gene encoding the methionine importer is a metP gene, and wherein the metP gene encodes a polypeptide having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of a polypeptide encoded by SEQ ID NOs:
- 1056, 1057, 1061, 1130 or 1132,
- a knock-out of an endogenous methionine efflux pump, wherein the endogenous methionine efflux pump is encoded by a yjeH gene,
- a phage deletion, and
- a ΔdapA auxotrophy.
A recombinant bacterial cell comprising:
- a heterologous methionine decarboxylase (metDC) gene operably linked to a promoter, wherein the heterologous metDC gene comprises a sequence that has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1133, 1134, or 1135,
- a heterologous gene encoding a methionine importer, wherein the heterologous gene encoding the methionine importer is a metP gene, and wherein the metP gene comprises a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1041, 1042, 1044, 1129 or 1131,
- a knock-out of an endogenous methionine efflux pump, wherein the endogenous methionine efflux pump is encoded by a yjeH gene,
- a phage deletion, and
- a ΔdapA auxotrophy.
The recombinant bacterial cell of any one of the previous claims, wherein the recombinant bacterial cell is a recombinant probiotic bacterial cell.
A pharmaceutical composition comprising the recombinant bacterial cell of any one of the previous claims and a pharmaceutically acceptable carrier.
The method of claim 52, wherein the increase is at least 1.2-fold, 1.5-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4-fold, 4.5-fold or 5-fold.
A non-naturally occurring MetDC enzyme comprising a sequence that has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to, comprises, or consists of any one of SEQ ID NOs:
- 1136, 1137, or 1138.
A host cell that comprises a heterologous polynucleotide encoding a methionine decarboxylase (MetDC) enzyme, wherein the heterologous polynucleotide comprises a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to, comprises, or consists of any one of SEQ ID NOs:
- 1133, 1134, or 1135.
A host cell that comprises a heterologous polynucleotide encoding a methionine decarboxylase (MetDC) enzyme, wherein the MetDC enzyme comprises an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to, comprises, or consists of a sequence selected from SEQ ID NOs:
- 1136, 1137, or 1138.
A host cell that comprises a heterologous polynucleotide encoding a methionine decarboxylase (MetDC) enzyme, wherein the sequence of the MetDC enzyme comprises one or more amino acid substitutions relative to the sequence of SEQ ID NO:
- 1049 at positions corresponding to position 41, 70, 82, 491 and/or 500 in SEQ ID NO:
- 1049.
A non-naturally occurring nucleic acid comprising a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to, comprises, or consists of any one of SEQ ID NOs:
- 1133, 1134, or 1135.
Dependencies are inferred from the claim text.