Recombinant Host Production of Consistent Rebaudioside D
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Solution Overview
Problem
Existing stevia extracts vary in composition and quality due to environmental factors, leading to inconsistent flavor profiles and the presence of contaminants, with rebaudioside D being a higher quality sweetener that is underrepresented in commercial preparations.
Innovation Solution
Development of recombinant hosts, such as microorganisms and plant cells, engineered to produce steviol glycosides like rebaudioside D through the expression of biosynthesis genes, particularly EUGT11 and other UDP glycosyl transferases, to enhance production and reduce plant-derived contaminants.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If natural stevia extracts are used, then sweetening function is provided, but composition consistency and quality are poor due to environmental factors and contaminants
Solution Approach 1:
The patent extracts only the beneficial glycosylation enzymes (UDP glycosyl transferases) from the Stevia plant and transfers them to recombinant host cells. This separates the desirable sweetening function from the harmful plant contaminants, allowing production of purified steviol glycosides without environmental variability or off-flavors.
Solution Approach 2:
The patent creates genetic copies of the EUGT11 and other UDP glycosyl transferase genes from Stevia rebaudiana and inserts them into recombinant host cells (such as yeast or bacteria). These copied enzymes replicate the plant's natural glycosylation capability while eliminating dependence on natural plant extraction, ensuring consistent composition and quality.
2Quantity of substance
If natural stevia extracts are used, then sweetening function is provided, but rebaudioside D levels are insufficient due to natural variation
Solution Approach 1:
The patent specifically enhances the expression of EUGT11, the enzyme responsible for producing rebaudioside D, in recombinant host cells. By locally optimizing this specific enzymatic pathway through genetic engineering, the system achieves high and stable rebaudioside D production while maintaining control over overall glycoside composition, eliminating natural variability.
3Reliability
If recombinant hosts are engineered to produce steviol glycosides, then production consistency and purity are improved, but manufacturing complexity increases
Solution Approach 1:
The patent uses universal recombinant host cell systems (such as yeast or E. coli) that can be engineered to express multiple Stevia-derived enzymes simultaneously. These hosts serve multiple functions: they provide the cellular machinery for enzyme production, enable scalable fermentation, and allow for consistent glycoside synthesis. This multi-functionality reduces overall manufacturing complexity despite the genetic engineering requirements.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The recombinant hosts produce steviol glycosides with enhanced levels of rebaudioside D, offering superior sweetening properties and reduced contaminants, addressing the variability and quality issues of natural stevia extracts.
Implementation Method 1
EUGT11, a uridine 5'-diphospho (UDP) glycosyl transferase described herein, can be used alone or in combination with one or more other UDP glycosyl transferases such as UGT74G1, UGT76G1, UGT85C2, and UGT91D2e, to allow the production and accumulation of rebaudioside D in recombinant hosts or using in vitro systems.
Data Source
AI summary
Recombinant microorganisms, plants, and plant cells are disclosed that have been engineered to express recombinant genes encoding UDP-glycosyltransferases (UGTs). Such microorganisms, plants, or plant cells can produce steviol glycosides, e.g., Rebaudioside A and/or Rebaudioside D, which can be used as natural sweeteners in food products and dietary supplements.