Protein with dextran-saccharase activity, and uses

Patent metadata

Field	Value
Publication number	US-10316300-B2
Application number	US-201515329941-A
Country	US
Kind code	B2
Filing date	Jul 21, 2015
Priority date	Jul 28, 2014
Publication date	Jun 11, 2019
Grant date	Jun 11, 2019

Abstract

The invention relates to: —a protein having dextran-saccharase activity and the sequence SEQ ID NO: 1 for an amino acid sequence; —a protein having dextran-saccharase activity and the sequence SEQ ID NO: 2 for an amino acid sequence; —a complex containing a substrate and a protein with dextran-saccharase activity —a method for synthesizing dextrans; and —dextrans. The invention also relates to a method for synthesizing gluco-oligosaccharides and gluco-conjugates and to the resulting products.

First claim

The invention claimed is:

1. A process for synthesizing dextrans comprising:
forming dextrans by contacting sucrose in a synthesis medium with
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1.

2. A process for synthesizing dextrans comprising:
forming dextrans by contacting sucrose in a synthesis medium with
(a) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1.

3. The process of claim 1 , wherein the step of contacting to form dextrans is carried out either at a temperature of between 20° C. and 50° C., or at a temperature of between 25° C. and 45° C.

4. The process of claim 1 , wherein the sucrose is present in the synthesis medium at either a concentration of between 50 and 600 g·l −1 , or a concentration of between 50 and 400 g·l −1 .

5. The process of claim 1 , wherein the step of contacting to form dextrans takes place either at a pH that is between 4 and 7, or at a pH of approximately 5.75.

6. A process for synthesizing glucooligosaccharides comprising:
forming glucooligosaccharides by contacting a synthesis medium that comprises sucrose and at least one carbohydrate acceptor with
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1.

7. A process for synthesizing glucoconjugate compound comprising:
forming glucoconjugate compounds by contacting synthesis medium that comprises sucrose and at least one hydroxylated molecule with
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1 and position 563 to 1282 within SEQ ID NO: 1.

8. A process for synthesizing glucooligosaccharides comprising:
(i) forming glucooligosaccharides by contacting a synthesis medium that comprises sucrose and at least one carbohydrate acceptor with
(a) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1.

9. A process for synthesizing glucoconjugate compounds comprising:
forming a glucoconjugate compounds by contacting a synthesis medium that comprises sucrose and at least one hydroxylated molecule with
(a) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 1 or
(b) a complex comprising a support and a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 1,
wherein said protein with dextransucrase activity has been immobilized on said support.

10. The process of claim 1 , wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

11. The process of claim 10 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

12. The process of claim 2 , wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

13. The process of claim 12 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

14. The process of claim 6 , wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

15. The process of claim 14 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

16. The process of claim 7 , wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

17. The process of claim 16 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

18. The process of claim 8 wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

19. The process of claim 18 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

20. The process of claim 9 , wherein the protein with dextransucrase activity is selected from
(a) a protein with dextransucrase activity where the protein is at least 95% identical to SEQ ID NO: 2 or
(b) a protein with dextransucrase activity where the protein is at least 98% identical to SEQ ID NO: 2.

21. The process of claim 20 , wherein the protein with dextransucrase activity comprises the amino acid sequence set forth in SEQ ID NO: 2.

Assignees

• Institut Nat Des Sciences Appliquees De Toulouse
• Agronomique Inst Nat Rech
• Centre Nat Rech Scient
• Univ Toulouse 3 Paul Sabatier
• Institut Nat Des Sciences Appliquees Toulouse

Inventors

• Vuillemin Marlène
• Claverie Marion
• Moulis Claire
• Remaud-Simeon Magali
• Monsan Pierre
• Severac Etienne
• Fontagne-Faucher Catherine

Classifications

• A61K8/73

  Polysaccharides · CPC title

• C12Y204/01005

  Dextransucrase (2.4.1.5) · CPC title

• A23L29/273

  Dextran; Polysaccharides produced by leuconostoc · CPC title

• C12P19/18

  produced by the action of a glycosyl transferase, e.g. alpha-, beta- or gamma-cyclodextrins · CPC title

• C12N9/1051 Primary

  Hexosyltransferases (2.4.1) · CPC title