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US 10,329,250 B2Granted
Process for producing 7-dehydrocholesterol and vitamin D3
AssigneeKyowa Hakko Bio Co., Ltd.
InventorsTetsuro Ujihara, Satoshi Mitsuhashi
Life of this patent
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Abstract
According to the present invention, there can be provided a process for producing 7-dehydrocholesterol (7DHC), comprising culturing, in a medium, a 7DHC-producing Labyrinthulea microorganism in which sterol 24-C-methyltransferase activity is reduced or lost as compared to a parent strain, allowing 7DHC to be produced and accumulated in the culture, and collecting the 7DHC from the culture; and a process for producing vitamin D3, comprising irradiating, with ultraviolet light, 7-dehydrocholesterol produced by the production process.
Claims6
A process for producing 7-dehydrocholesterol (hereinafter “7DHC”), comprising:
- culturing, in a medium, a 7DHC-producing Labyrinthulea microorganism in which sterol 24-C-methyltransferase activity (hereinafter “SMT activity”) is reduced or lost as compared to a parent strain;
- allowing 7DHC to be produced and accumulated in the culture;
- and
- collecting the 7DHC from the culture,
- wherein the Labyrinthulea microorganism in which the SMT activity is reduced or lost as compared to the parent strain is a Labyrinthulea microorganism in which the SMT activity is reduced or lost as compared to the parent strain through deletion, substitution, or addition of at least one base in a gene which is present in the chromosomal DNA of the parent strain and encodes a protein having SMT activity, and
- wherein the gene encoding a protein having SMT activity is a gene having any of the following DNAs [1] to [5]:
- [1] a DNA encoding a protein having the amino acid sequence shown by SEQ ID NO:
- 2;
- [2] a DNA encoding a homologous protein having at least 95% identity with the amino acid sequence shown by SEQ ID NO:
- 2, and having SMT activity;
- [3] a DNA having the base sequence shown by SEQ ID NO:
- 1;
- [4] a DNA that hybridizes with a DNA consisting of a base sequence complementary to the base sequence shown by SEQ ID NO:
- 1 under stringent conditions, and encodes a homologous protein having SMT activity, wherein the stringent conditions comprise conditions in which a filter on which a DNA has been immobilized and a probe DNA are incubated overnight at 42° C. in a solution containing 50% formamide, 5×SSC (750 mM sodium chloride and 75 mM sodium citrate), 50 mM sodium phosphate (pH 7.6), 5×Denhardt's solution, 10% dextran sulfate, and 20 μg/l of a denatured salmon sperm DNA, and then the filter is washed in a 0.2×SSC solution at about 65° C.;
- and
- [5] a DNA having at least 95% identity with the base sequence shown by SEQ ID NO:
- 1, and encoding a homologous protein having SMT activity.
14 more elements
A process for producing 7-dehydrocholesterol (hereinafter “7DHC”), comprising:
- culturing, in a medium, a 7DHC-producing Labyrinthulea microorganism in which sterol 24-C-methyltransferase activity (hereinafter “SMT activity”) is reduced or lost as compared to a parent strain;
- allowing 7DHC to be produced and accumulated in the culture;
- and
- collecting the 7DHC from the culture,
- wherein the Labyrinthulea microorganism in which the SMT activity is reduced or lost as compared to the parent strain is a Labyrinthulea microorganism in which the SMT activity is reduced or lost as compared to the parent strain, being obtained by transforming the parent strain with a DNA encoding an RNA which has a sequence complementary to a messenger RNA transcribed from a gene present in the chromosomal DNA of the parent strain and encoding a protein having SMT activity, and forms a conjugate with the messenger RNA, and
- wherein the gene encoding a protein having SMT activity is a gene having any of the following DNAs [1] to [5]:
- [1] a DNA encoding a protein having the amino acid sequence shown by SEQ ID NO:
- 2;
- [2] a DNA encoding a homologous protein having at least 95% identity with the amino acid sequence shown by SEQ ID NO:
- 2, and having SMT activity;
- [3] a DNA having the base sequence shown by SEQ ID NO:
- 1;
- [4] a DNA that hybridizes with a DNA consisting of a base sequence complementary to the base sequence shown by SEQ ID NO:
- 1 under stringent conditions, and encodes a homologous protein having SMT activity, wherein the stringent conditions comprise conditions in which a filter on which a DNA has been immobilized and a probe DNA are incubated overnight at 42° C. in a solution containing 50% formamide, 5×SSC (750 mM sodium chloride and 75 mM sodium citrate), 50 mM sodium phosphate (pH 7.6), 5×Denhardt's solution, 10% dextran sulfate, and 20 μg/l of a denatured salmon sperm DNA, and then the filter is washed in a 0.2×SSC solution at about 65° C.;
- and
- [5] a DNA having at least 95% identity with the base sequence shown by SEQ ID NO:
- 1, and encoding a homologous protein having SMT activity.
14 more elements
Dependencies are inferred from the claim text.