METHOD FOR THE PREPARATION OF 2,4-DIHYDROXYBUTYRATE
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Abstract
A method for the preparation of 2,4-dihydroxybutyric acid from homoserine includes a first step of conversion of the primary amino group of homoserine to a carbonyl group to obtain 2-oxo-4-hydroxybutyrate, and a second step of reduction of the obtained 2-oxo-4-hydroxybutyrate (OHB) to 2,4-dihydroxybutyrate.
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39 Claims
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1-22. -22. (canceled)
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23. A method for the preparation of 2,4-dihydroxybutyrate (2,4-DHB) from homoserine comprising:
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deaminating homoserine to form 2-oxo-4-hydroxybutyrate (OHB), where the deamination of homoserine is catalyzed by an enzyme having homoserine transaminase activity, wherein the enzyme having homoserine transaminase activity is produced via a transformed host microorganism that comprises a first chimeric gene including a first nucleic acid sequence encoding the enzyme having homoserine transaminase activity for converting the primary amino acid group of homoserine to a carbonyl group to obtain OHB; and reducing the OHB to form 2,4-DHB, where the reduction of OHB is catalyzed by an enzyme having OHB reductase activity, wherein the enzyme having OHB reductase activity is produced via the transformed host microorganism, which further comprises a second chimeric gene including a second nucleic acid sequence encoding the enzyme having OHB reductase activity for reducing OHB to 2,4-DHB. - View Dependent Claims (24, 25, 26, 27, 28, 29, 30)
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31. A modified microorganism for the preparation of 2,4-dihydroxybutyrate (2,4-DHB) from homoserine via a two-step pathway comprising:
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deaminating homoserine to form 2-oxo-4-hydroxybutyrate (OHB), where the deamination of homoserine is catalyzed by an enzyme having homoserine transaminase activity, and reducing the OHB to form 2,4-DHB, where the reduction of OHB is catalyzed by an enzyme having OHB reductase activity; wherein the modified microorganism is a host microorganism that has been transformed to enhance production of 2,4-DHB compared to a non-transformed host microorganism, the transformed host microorganism comprising; a first chimeric gene including a first nucleic acid sequence encoding the enzyme having homoserine transaminase activity for converting the primary amino acid group of homoserine to a carbonyl group to obtain OHB, and a second chimeric gene including a second nucleic acid sequence encoding the enzyme having OHB reductase activity for reducing OHB in 2,4-DHB. - View Dependent Claims (32, 33, 34, 35, 36, 37, 38, 39)
has at least one of the genes deleted chosen among IdhA, adhE, ackA, pta, poxB, focA, pfIB, sad, gabABC, sfcA, maeB, ppc, pykA, pykF, mgsA, sucAB, ptsl, ptsG, pgi, fumABCaldA, HdD, icIR, metA, thrB, lysA, eda, rthA, rthB, and rthC.
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37. The modified microorganism of claim 31, wherein the enzyme having homoserine transaminase activity is selected from the group consisting of enzymes classified in E.C. 2.6.1.1, E.C. 2.6.1.2, E.C. 2.6.1.42, E.C. 2.6.1.57 or E.C. 2.6.1.88, and/or wherein the enzyme having OHB reductase activity is a lactate dehydrogenase classified in E.C.1.1.1.27 or E.C.1.1.1.28, a malate dehydrogenase classified in E.C.1.1.1.37, E.C.1.1.1.82 or E.C.1.1.1.299, or a branched-chain 2-hydroxyacid dehydrogenase classified in E.C.1.1.1.272 or E.C.1.1.1.345.
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38. A method of production of 2,4-DHB comprising the steps of
culturing the modified microorganism of claim 31 in an appropriate culture medium, recovering 2,4-DHB from the culture medium. -
39. The method of claim 38 wherein the 2,4-DHB is further purified.
Specification