Biosynthesis of phloroglucinol and preparation of 1,3-dihydroxybenzene therefrom
AssigneeBoard of Trustees of Michigan State University
InventorJohn Frost
Life of this patent
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Abstract
The present invention provides methods, enzymes, and cells for the biosynthetic production of phloroglucinol from malonyl-CoA, which is ultimately obtained from simple starting materials such as glucose; also provided are methods for preparing derivatives of biosynthetic phloroglucinol, including, e.g., resorcinol.
Claims60
An isolated or recombinant PhlD+ enzyme system that is PhlA−, PhlB−, and PhlC−, or a PhlD+ recombinant cell that is PhlA−, PhlB−, and PhlC−, or a PhlD+, PhlA−, PhlB−, and PhlC− recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein, the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol.
A process for production of anabolic phloroglucinol, comprising the steps of
- (A) providing
- (1) (a) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−, or
- (b) a PhlD+ recombinant cell, that is at least one of PhlA−, PhlB−, or PhlC−, or
- (c) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein,
- the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (2) at least one of malonyl-CoA or another carbon source that the enzyme system or recombinant cell is capable of converting to malonyl-CoA;
- and
- (B) contacting
- (1) the malonyl-CoA with the enzyme system or recombinant cell under conditions in which it can synthesize phloroglucinol therefrom, or
- (2) the other carbon source with the enzyme system or recombinant cell under conditions in which it can convert the carbon source to malonyl-CoA and can synthesize phloroglucinol therefrom;
- thereby obtaining anabolic phloroglucinol.
Isolated anabolic phloroglucinol.
An isolated or recombinant phloroglucinol synthase enzyme capable of converting malonyl-CoA to phloroglucinol, wherein said enzyme comprises an amino acid sequence that is at least 90% homologous to SEQ ID NO:2.
Isolated or recombinant nucleic acid vector comprising at least one open reading frame encoding a PhlD enzyme, and that is at least one of phlA−, phlB−, or phlC−, and which further comprises at least one open reading frame encoding a malonyl-CoA synthesis enzyme.
A phlD+ recombinant cell, wherein said cell has been transformed with a nucleic acid vector comprising at least one open reading frame encoding a PhlD enzyme, from which the cell can express phloroglucinol synthase, the recombinant cell being phlA−, phlB−, and phlC− and being capable of converting malonyl-CoA to phloroglucinol by action of phloroglucinol synthase expressed therein.
A process for the preparation of resorcinol, comprising
- (A)(1) anabolically biosynthesizing phloroglucinol by action of (a) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−, or (b) a PhlD+ recombinant cell that is at least one of PhlA−, PhlB−, or PhlC−, or (c) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein, the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (2) providing hydrogen and a rhodium catalyst;
- and
- (B) contacting phloroglucinol biosynthesized thereby with the hydrogen and the rhodium catalyst under conditions in which the phloroglucinol is hydrogenated to form resorcinol,
- thereby preparing resorcinol.
A method for manufacturing a medicament, cosmetic, dye, polymer resin, rubber, adhesive, sealant, coating, composite material, or laminated or bonded material, comprising
- (1) anabolically biosynthesizing phloroglucinol by action of (a) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−, or (b) a PhlD+ recombinant cell that is at least one of PhlA−, PhlB−, or PhlC−, or (c) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein, the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (2) preparing a medicament, cosmetic, dye, polymer resin, rubber, adhesive, sealant, coating, composite material, or laminated or bonded material from anabolic phloroglucinol biosynthesized thereby, or from resorcinol obtained by chemically modifying the anabolic phloroglucinol.
A medicament, cosmetic, dye, polymer resin, rubber, adhesive, sealant, coating, composite material, or laminated or bonded material prepared by a process comprising
- (1) anabolically biosynthesizing phloroglucinol by action of (a) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−;
- or (b) a PhlD+ recombinant cell that is at least one of PhlA−, PhlB−, or PhlC−;
- or (c) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein;
- the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (2) preparing a medicament, cosmetic, dye, polymer resin, rubber, adhesive, sealant, coating, composite material, or laminated or bonded material from anabolic phloroglucinol biosynthesized thereby, or from resorcinol obtained by chemically modifying the anabolic phloroglucinol.
A process for the preparation of a recombinant cell that is capable of biosynthesizing phloroglucinol from malonyl-CoA, comprising
- (A) transforming a cell with a PhlD-encoding nucleic acid by
- (1) providing isolated or recombinant nucleic acid that is phlA−, phlB−, and phlC− and that contains at least one open reading frame encoding a PhlD enzyme, or contains both at least one open reading frame encoding a PhlD enzyme and at least one open reading frame encoding a malonyl-CoA synthesis enzyme,
- (2) providing (a) a PhlD−, phlA−, phlB−, and phlC− cell that can be transformed with said nucleic acid, or (b) a PhlD+, phlA−, phlB−, and phlC− cell that can be transformed with said nucleic acid;
- and
- (3) contacting the cell with the nucleic acid under conditions in which the cell can be transformed with the nucleic acid,
- the nucleic acid being capable of expression by the cell, thereby obtaining a recombinant cell;
- or
- (B) inactivating genes in a phlD+ cell by
- (1) providing a phlD+ cell that is at least one of phlA+, phlB+, or phlC+;
- and
- (2) inactivating all of said phlA, phlB, and phlC genes therein,
- thereby obtaining a recombinant cell that is phlA−, phlB−, and phlC−.
A method for producing a propellant or explosive compound comprising
- (A) anabolically biosynthesizing phloroglucinol by action of (1) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−, or (2) a PhlD+ recombinant cell that is at least one of PhlA−, PhlB−, or PhlC−, or (3) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein, the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (B) chemically modifying anabolic phloroglucinol biosynthesized thereby, or chemically modifying resorcinol prepared therefrom, to produce a propellant or explosive compound.
An explosive or propellant composition prepared by a process comprising
- (1) anabolically biosynthesizing phloroglucinol by action of (a) an isolated or recombinant PhlD+ enzyme system that is at least one of PhlA−, PhlB−, or PhlC−;
- or (b) a PhlD+ recombinant cell that is at least one of PhlA−, PhlB−, or PhlC−;
- or (c) a PhlD+ recombinant cell that is a PhlD+ cell that has been genetically engineered to further increase the expression of PhlD therein;
- the enzyme system or recombinant cell being capable of converting malonyl-CoA to phloroglucinol;
- and
- (2) preparing an explosive or propellant composition from anabolic phloroglucinol biosynthesized thereby, or from resorcinol obtained by chemically modifying the anabolic phloroglucinol.
A composition comprising isolated anabolic phloroglucinol or a derivative thereof.
Dependencies are inferred from the claim text.