ENZYMATIC PRODUCTION OF D-TAGATOSE

• US 20200199637A1
• Filed: 01/07/2020
• Published: 06/25/2020
• Est. Priority Date: 10/02/2015
• Status: Active Grant

• Current Assignee: Bonumose LLC
• Original Assignee: Bonumose LLC
• Inventors: WICHELECKI, Daniel Joseph
• Granted Patent: US 11,034,988 B2
• Time in Patent Office: Days
• CPC Class Codes: C12N 9/90 Isomerases (5.)
• CPC Class Codes: C12P 19/02 Monosaccharides 2-ketogulonic acid C12P7/60
• CPC Class Codes: C12Y 207/01144 Tagatose-6-phosphate kinase (2.7.1.144)

Abstract

The current disclosure provides a process for enzymatically converting a saccharide into tagatose. The invention also relates to a process for preparing tagatose where the process involves converting fructose 6-phosphate (F6P) to tagatose 6-phosphate (T6P), catalyzed by an epimerase, and converting the T6P to tagatose, catalyzed by a phosphatase.

Claims

1. A process for preparing tagatose, the process comprising:

converting fructose 6-phosphate (F6P) to tagatose 6-phosphate (T6P) catalyzed by an epimerase; and

converting the T6P produced to tagatose catalyzed by a phosphatase.

2. The process of claim 1, further comprising a step of converting glucose 6-phosphate (G6P) to the F6P, wherein the step is catalyzed by phosphoglucose isomerase (PGI).

3. The process of claim 2, further comprising the step of converting glucose 1-phosphate (G1P) to the G6P, wherein the step is catalyzed by phosphoglucomutase (PGM).

4. The process of claim 3, further comprising the step of converting a saccharide to the G1P, wherein the step is catalyzed by at least one enzyme, wherein the saccharide is selected from the group consisting of a starch or derivative thereof, cellulose or a derivative thereof and sucrose.

5. The process of claim 4, wherein at least one enzyme is selected from the group consisting of alpha-glucan phosphorylase (α

GP), maltose phosphorylase, sucrose phosphorylase, cellodextrin phosphorylase, cellobiose phosphorylase, and cellulose phosphorylase.

7. The process of claim 4, further comprising the step of converting starch to a starch derivative wherein the starch derivative is prepared by enzymatic hydrolysis of starch or by acid hydrolysis of starch.

9. The process of claim 7, wherein the starch derivative is prepared by enzymatic hydrolysis of starch catalyzed by isoamylase, pullulanase, alpha-amylase, or a combination thereof.

10. The process of claim 1, further comprising:

the step of converting fructose to the F6P, wherein the step is catalyzed by at least one enzyme; and

optionally, the step of converting sucrose to the fructose, wherein the step is catalyzed by at least one enzyme.

11. The process of claim 2, further comprising:

the step of converting glucose to the G6P, wherein the step is catalyzed by at least one enzyme, and

optionally, the step of converting sucrose to the glucose, wherein the step is catalyzed by at least one enzyme.

12. The process of claim 1, wherein the epimerase is fructose 6-phosphate epimerase.

13. The process of claim 12, wherein the fructose 6-phosphate epimerase is encoded by a polynucleotide comprising a nucleotide sequence having at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% sequence identity with SEQ ID NOS.:

1, 3, 5, 7, 9, or 10.

14. The process of claim 12, wherein the fructose 6-phosphate epimerase comprises an amino acid sequence having at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% sequence identity with SEQ ID NOS.:

2, 4, 6, 8, or 11.

15. The process of claim 1, wherein the phosphatase is tagatose 6-phosphate phosphatase.

16. The process of claim 15, wherein the tagatose 6-phosphate phosphatase is encoded by a polynucleotide comprising a nucleotide sequence having at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% sequence identity with SEQ ID NOS.:

12, 14, or 16.

17. The process of claim 15, wherein the tagatose 6-phosphate phosphatase comprises an amino acid sequence having at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or 100% sequence identity with SEQ ID NOS.:

13, 15, or 17.

18. The process of claim 117, wherein the process steps are conducted at a temperature ranging from about 40°

C. to about 70°

C., at a pH ranging from about 5.0 to about 8.0, and/or for about 8 hours to about 48 hours.

19. The process of claim 118, wherein the process steps are conducted in one bioreactor or in a plurality of bioreactors arranged in series.

20. The process of claim 1, wherein the process steps are conducted ATP-free, NAD(H)-free, at a phosphate concentration from about 0 mM to about 150 mM, the phosphate is recycled, and/or at least one step of the process involves an energetically favorable chemical reaction.

21. Tagatose prepared by a process of claim 1.

6. (canceled)

8. The process of claim 6, wherein 4-glucan transferase (4GT) is added to the process.