Transformed Synechococcus elongatus strain having capability of producing squalene from carbon dioxide and method for producing squalene using the same

Patent metadata

Field	Value
Publication number	US-10155964-B2
Application number	US-201615357815-A
Country	US
Kind code	B2
Filing date	Nov 21, 2016
Priority date	Apr 5, 2016
Publication date	Dec 18, 2018
Grant date	Dec 18, 2018

Abstract

The present specification discloses a transformed Synechococcus elongatus strain which may directly produce squalene from carbon dioxide, and a method for producing squalene and a method for removing carbon dioxide, using the same. In an aspect, the strain may produce squalene using carbon dioxide as a carbon source. The Synechococcus elongatus strain is economically efficient because a high-value added squalene is produced using light and carbon dioxide present in the atmosphere as a carbon source, and the method for producing squalene is eco-friendly because the strain may be utilized to remove or reduce carbon dioxide in the atmosphere by using microorganisms. The strain of the present disclosure may produce only squalene, which is a desired target material with high purity, and has an advantage in that squalene may be continuously mass-produced.

First claim

What is claimed is:

1. A Synechococcus elongatus strain comprising:
a gene encoding an enzyme producing 1-deoxy-D-xylulose 5-phosphate (DXP) from pyruvate and D-glyceraldehyde 3-phosphate (G3P) comprising a sequence of SEQ ID NO. 1;
a gene encoding an enzyme producing dimethylallyl diphosphate (DMAPP) from isopentenyl diphosphate (IPP) comprising a sequence of SEQ ID NO. 2;
a gene encoding an enzyme producing farnesyl diphosphate (FPP) from dimethylallyl diphosphate (DMAPP) comprising a sequence of SEQ ID NO. 3; and
a gene encoding an enzyme producing squalene from farnesyl diphosphate (FPP) comprising a sequence of SEQ ID NO. 4 or 5.

2. The Synechococcus elongatus strain according to claim 1 , wherein the enzyme producing 1-deoxy-D-xylulose 5-phosphate (DXP) from pyruvate and D-glyceraldehyde 3-phosphate (G3P) is a deoxyxylulose-5-phosphate synthase,
the enzyme producing dimethylallyl diphosphate (DMAPP) from isopentenyl diphosphate (IPP) is an isopentenyl diphosphate delta isomerase,
the enzyme producing farnesyl diphosphate (FPP) from dimethylallyl diphosphate (DMAPP) is a geranyl diphosphate synthase, and
the enzyme producing squalene from farnesyl diphosphate (FPP) is a squalene synthase.

3. The Synechococcus elongatus strain according to claim 1 , wherein the gene encoding the enzyme producing 1-deoxy-D-xylulose 5-phosphate (DXP) from pyruvate and D-glyceraldehyde 3-phosphate (G3P);
the gene encoding the enzyme producing dimethylallyl diphosphate (DMAPP) from isopentenyl diphosphate (IPP); and
the gene encoding the enzyme producing farnesyl diphosphate (FPP) from dimethylallyl diphosphate (DMAPP) are each derived from E. coli , and
the gene encoding the enzyme producing squalene from farnesyl diphosphate (FPP) is derived from Saccharomyces cerevisiae or Methylococcus capsulatus.

4. The Synechococcus elongatus strain according to claim 1 , wherein the strain is transformed with a first vector and a second vector,
the first vector comprising:
a gene encoding an enzyme producing 1-deoxy-D-xylulose 5-phosphate (DXP) from pyruvate and D-glyceraldehyde 3-phosphate (G3P);
a gene encoding an enzyme producing dimethylallyl diphosphate (DMAPP) from isopentenyl diphosphate (IPP); and
a gene encoding an enzyme producing farnesyl diphosphate (FPP) from dimethylallyl diphosphate (DMAPP), and
the second vector comprising: a gene encoding an enzyme producing squalene from farnesyl diphosphate (FPP).

5. The Synechococcus elongatus strain according to claim 4 , wherein the first vector comprises a sequence of SEQ ID NO. 7.

6. The Synechococcus elongatus strain according to claim 4 , wherein the second vector comprises a sequence of SEQ ID NO. 9, or a sequence of SEQ ID NO. 10.

7. The Synechococcus elongatus strain according to claim 4 , wherein the first vector is inserted into a neutral site I (NSI) of Synechococcus elongatus which is a parent strain, and the second vector is inserted into a neutral site II (NSII) of Synechococcus elongatus which is a parent strain.

8. The Synechococcus elongatus strain according to claim 4 , wherein the first vector sequentially comprises:
a spectinomycin-resistant gene;
a lacI repressor;
a trc promoter; and
a target gene(s), and
wherein the target genes are
the gene encoding the enzyme producing 1-deoxy-D-xylulose 5-phosphate (DXP) from pyruvate and D-glyceraldehyde 3-phosphate (G3P);
the gene encoding the enzyme producing dimethylallyl diphosphate (DMAPP) from isopentenyl diphosphate (IPP); and
the gene encoding the enzyme producing farnesyl diphosphate (FPP) from dimethylallyl diphosphate (DMAPP).

9. The Synechococcus elongatus strain according to claim 4 , wherein the second vector sequentially comprises:
a kanamycin-resistant gene;
a lacI repressor;
a trc promoter; and
a target gene encoding the enzyme producing squalene from farnesyl diphosphate (FPP).

10. The Synechococcus elongatus strain according to claim 4 , wherein the strain is a strain in which the vectors are transformed in Synechococcus elongatus PCC7942 (Accession number: ATCC 33912), which is a parent strain.

11. The Synechococcus elongatus strain according to claim 1 , wherein the strain is a strain belonging to accession number KCTC 12966BP.

Assignees

• Korea Inst Sci & Tech

Inventors

• Woo Han Min
• Choi Sun-Young
• Um Youngsoon
• Gong Gyeong Taek
• Lee Sun Mi
• Kim Yunje

Classifications

• C12P5/007 Primary

  containing one or more isoprene units, i.e. terpenes (carotenes C12P23/00) · CPC title

• C12Y205/01021

  Squalene synthase (2.5.1.21), i.e. farnesyl-disphosphate farnesyltransferase · CPC title

• C12N9/1085

  transferring alkyl or aryl groups other than methyl groups (2.5) · CPC title

• C12Y101/01267

  1-Deoxy-D-xylulose-5-phosphate reductoisomerase (1.1.1.267) · CPC title

• C12N15/52

  Genes encoding for enzymes or proenzymes · CPC title