Enzymatic production of mannose
Disclosed herein are improved processes for making mannose including the step of converting the M6P to mannose, catalyzed by a M6PP, using enzymes with higher activities compared to M6PPs previously used in a process to produce mannose.
1. An improved enzymatic process for the production of mannose from a saccharide, the improvement comprising converting mannose 6-phosphate (M6P) to mannose catalyzed by a mannose-6-phosphate phosphatase (M6PP), wherein the M6PP comprises the amino acid sequence as set forth in SEQ ID NO: 1,
wherein the mannose yield is at least three-fold more than the mannose yield produced by catalyzing the conversion of M6P to mannose catalyzed by an M6PP from Sulfurivirga caldicuralii under equivalent process conditions.
2. The process of claim 1 , further comprising a step of converting fructose-6-phosphate (F6P) to mannose 6-phosphate (M6P) catalyzed by a mannose 6-phosphate isomerase (M6PI).
3. The process of claim 2 , further comprising a step of converting glucose 6-phosphate (G6P) to the F6P, wherein the step is catalyzed by a phosphoglucose isomerase (PGI).
4. The process of claim 3 , further comprising a step of converting glucose 1-phosphate (G1P) to the G6P, wherein the step is catalyzed by a phosphoglucomutase (PGM).
5. The process of claim 4 , further comprising a 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, a starch derivative, cellulose, a cellulose derivative, and sucrose.
6. The process of claim 5 , wherein the at least one enzyme is selected from the group consisting of alpha-glucan phosphorylase (aGP), maltose phosphorylase, sucrose phosphorylase, cellodextrin phosphorylase, cellobiose phosphorylase, and cellulose phosphorylase.
7. The process of claim 5 , wherein the saccharide is starch or a derivative thereof selected from the group consisting of amylose, amylopectin, soluble starch, amylodextrin, maltodextrin, maltose, and glucose.
8. The process of claim 7 , further comprising a 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 8 , further comprising a step transglycosylating the starch derivative, catalyzed by a 4-glucan transferase (4GT).
10. The process of claim 9 , wherein the starch derivative is prepared by enzymatic hydrolysis of starch catalyzed by an isoamylase, a pullulanase, an alpha-amylase, or a combination thereof.
11. The process claim 2 , further comprising:
a step of converting fructose to the F6P, wherein the step is catalyzed by at least one enzyme; and, optionally, a step of converting sucrose to the fructose, wherein the step is catalyzed by at least one enzyme.
12. The process of claim 3 , further comprising:
a step of converting glucose to the G6P, wherein the step is catalyzed by at least one enzyme; and, optionally, a step of converting sucrose to the glucose, wherein the step is catalyzed by at least one enzyme.
13. The process of claim 1 , wherein the conversion of M6P to mannose is preceded by the steps of:
(i) converting a saccharide to glucose 1-phosphate (G1P) catalyzed by one or more enzymes, wherein the saccharide is selected from the group consisting of starch, one or more derivatives of starch, or a combination thereof;
(ii) converting G1P to glucose 6-phosphate (G6P) catalyzed by a phosphoglucomutase (PGM);
(iii) converting G6P to fructose 6-phosphate (F6P) catalyzed by a phosphoglucoisomerase (PGI); and
(iv) converting the F6P to mannose 6-phosphate (M6P) catalyzed by a mannose 6-phosphate isomerase (M6PI),
wherein all process steps are conducted as a continuous cascade reaction in a single reaction vessel.
14. The process of claim 13 , wherein the continuous cascade reaction process steps are conducted under at least one of the following process conditions:
at a temperature ranging from 37° C. to 85° C.,
at a pH ranging from 5.0 to 9.0, or
for 1 hour to 48 hours.
15. The process of claim 13 , wherein the continuous cascade reaction process steps are conducted under at least one of the following process conditions:
without adenosine triphosphate (ATP) as a source of phosphate,
without nicotinamide adenosine dinucleotide,
at a phosphate concentration from 0.1 mM to 150 mM,
at a Mg 2+ concentration from 0.1 mM to 50 mM
where phosphate is recycled, and
where at least one step of the process involves an energetically favorable chemical reaction.
16. The process of claim 15 , wherein phosphate ions produced by M6PP dephosphorylation of M6P are recycled in the process step of converting a saccharide to G1P.
17. The process of claim 15 , wherein the step of converting M6P to mannose is an energetically favorable, irreversible phosphatase reaction.
18. The process of claim 13 , wherein the derivatives of starch are selected from the group consisting of amylose, amylopectin, soluble starch, amylodextrin, maltodextrin, maltotriose, maltose, and glucose.