Production of Biofuels, Biomaterials and Sugar Alcohols (Polyols) by a Recombinant Microorganism Expressing Enzymes (Det1 and Pho13) for NADPH Upregulation and Polyol Phosphatase (Ynl010w) for Polyol Phosphatase Dephosphorylation

Description:

A New Enzyme and Process for Metabolic Engineering of Production
of Sorbitol, Mannitol, Ribitol, Xylitol, Arabitol, Erythritol, and Other Polyols

 

Princeton Docket # 13-2939

 

Scientists in the Lewis-Sigler Institute, Princeton University have discovered an enzyme which encodes a broad-spectrum, D-stereoisomer-specific sugar alcohol phosphatase, which has high activity against sorbitol-6-phosphate, mannitol-6-phosphate, ribitol-5-phosphate, xylitol-5-phosphate, arabitol-5-phosphate, and erythritol-4-phosphate. This is the first known enzyme to catalyze specific hydrolysis of these sugar alcohol phosphates. Over-expression of this enzyme, together with a sugar phosphate dehydrogenase (which converts sugar phosphate to polyol phosphate), represents a novel engineering pathway for polyol production. In combination with engineering to direct the carbon flow in central metabolism toward the production of the corresponding sugar phosphate, this approach can be applied to the production of many D-polyols. In addition, this enzyme could be utilized to clean up unnecessary and inhibitory polyol phosphates which could be generated in many other metabolic engineering processes where cells possess high sugar phosphate pools under a reducing environment. 

Applications:   

 

·         Production of sorbitol, mannitol, ribitol, xylitol, arabitol, or erythritol

 

·         These and other polyols (sugar alcohols) are useful as biomaterials and food, flavor, and pharmaceutical components

Advantages:

             

·         Improved  yields and productivity

 

·         Improved product separation

 

·         Opportunity to produce bulk quantities of novel polyols

Background

The bioproduction of polyols, a family of sugar alcohols, has drawn wide interest. Polyols are sweet like sugar, and share many of sugar’s physical properties, but are low calorie and do not contribute to high blood sugar levels or tooth decay. Thus, they are valuable sugar substitutes. Polyols are also broadly used in food, cosmetics, and pharmaceutics.  For example, sorbitol is commercially used as a laxative, humectant, thickener, and cryoprotectant. In addition, polyols are precursors to alkanes.

 

Polyols are naturally produced by plants and fungi at low levels. To achieve a high yield of polyols, the standard engineering approach is to first dephosphorylate the sugar phosphate and then reduce the resulting sugar to the polyol. Disadvantages of this approach are that the dephosphorylated sugar may be lost to other pathways, and that the reduction step may be non-specific. The newly discovered sugar alcohol phosphatase opens an alternative pathway, where dephosphorylation desirably leads directly to the polyol product. The novel pathway to D-polyols mirrors the natural glycerol excretion pathway, where sn-(L)-glycerol-3-phosphate is made from the reduction of dihydroxyacetone phosphate, followed by the dephosphorylation of the phosphorylated L-polyol.

 

Polyol phosphates, at high concentration, are also known to be inhibitory to primary metabolism because they structurally mimic the enediol intermediate of key enzymes (phosphoglucose isomerase, ribose-phosphate isomerase, triose-phosphate isomerase). Polyol phosphates are highly produced in many metabolic engineering processes where highly concentrated sugar phosphates are reduced to polyol phosphates. In plants, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCo) also has an enediol intermediate and is also known to be inhibited by a polyol phosphate, 2-carboxy-D-arabitol-1-phosphate. The newly discovered sugar alcohol phosphatase provides a powerful way to clean up these undesired polyol phosphates, which could potentially result in a higher central carbon metabolic flux and better yield in both microbes and plants.

 

Faculty Inventor: Dr. Joshua Rabinowitz, M.D., Ph.D.

Dr. Rabinowitz is a Professor at the Department of Chemistry and Lewis-Sigler Institute for Integrative Genomics at Princeton University. Dr. Rabinowitz has expertise in applying metabolomic technology to identify novel enzymes, drug targets, ... and metabolic pathways and regulatory mechanisms. Dr. Rabinowitz is an inventor of more than 130 U.S. Patents and author of over 110 major journal articles, including seven in the past four years in Science or Nature. He is co-founder of Alexza Pharmaceuticals, a member of the Kadmon Pharmaceuticals Scientific Advisory Board, and consultant to several major pharmaceutical and metabolic engineering companies. Dr. Rabinowitz earned two B.A. degrees, in Mathematics and Chemistry, from University of North Carolina at Chapel Hill, and his Ph.D. in Biophysics and his M.D. from Stanford University.

Publications

Pending and available under confidentiality.

Intellectual Property Status

Patent protection is pending.  Princeton is currently seeking partners for the further development and commercialization of this technology.

Contact

Laurie Tzodikov
Princeton University Office of Technology Licensing • (609) 258-7256•
tzodikov@princeton.edu

Princeton # 13-2939

 

 

 

 

Patent Information:
For Information, Contact:
Laurie Tzodikov
Licensing Associates
Princeton University
tzodikov@princeton.edu
Inventors:
YI-Fan Xu
Joshua Rabinowitz
Fabien Letisse
Sarah Johnson
Keywords:
biofuels
Biotechnology/Pharmaceuticals
Chemistry
metabolic engineering
model organism
yeast