Enzymatic Fructose-6-Phosphate Synthesis Bypassing Glucose Inhibition

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Solution Overview

Problem

Biological processes for producing fructose-6-phosphate from dihydroxyacetone phosphate and glyceraldehyde-3-phosphate are hindered by high glucose or sucrose concentrations, which inhibit enzymes like fructose bisphosphatase, reducing efficiency in fermentation processes.

Innovation Solution

A method involving the enzymatic conversion of dihydroxyacetone phosphate to dihydroxyacetone and subsequent condensation with glyceraldehyde-3-phosphate to produce fructose-6-phosphate, bypassing enzyme regulation by glucose or sucrose, using enzymes such as sugar phosphatase and fructose-6-phosphate aldolase.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If conventional enzymatic conversion using fructose bisphosphatase is used to produce fructose-6-phosphate, then the conversion can proceed through standard metabolic pathways, but the process is inhibited by high glucose or sucrose concentrations

Engineering Contradiction:

Improveconversion efficiencyVSAvoidenzyme inhibition by glucose/sucrose

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The conventional single-step enzymatic conversion is segmented into multiple sequential steps, each catalyzed by a different enzyme. The pathway is divided into: (1) DHAP dephosphorylation to DHA by phosphatase, (2) DHA condensation with G3P to form F6P by aldolase, or alternatively (3) G3P dephosphorylation to glyceraldehyde, (4) glyceraldehyde condensation with DHAP to form F1P by aldolase, and (5) F1P isomerization to F6P by phosphoglucomutase. This segmentation allows bypassing the inhibited fructose bisphosphatase step.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

Dihydroxyacetone (DHA) and/or glyceraldehyde serve as intermediary compounds that mediate the conversion from phosphate esters to fructose-6-phosphate. These intermediaries allow the reaction to proceed through alternative enzymatic pathways that are not subject to glucose/sucrose inhibition, effectively acting as mediators that circumvent the harmful inhibition effect.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Ease of manufacture

If the standard pathway through fructose-1,6-bisphosphate is used, then the conversion follows central carbon metabolism, but enzyme regulation by glucose or sucrose reduces process efficiency

Engineering Contradiction:

Improveprocess simplicityVSAvoidfermentation efficiency

Core Design Contradiction:

Ease of manufactureVSProductivity

Solution Approach 1:

Instead of following the conventional forward direction of gluconeogenesis (F6P → F1,6BP → DHAP + G3P), the invention inverts the pathway by condensing DHAP and G3P to form F6P directly through alternative enzymes. This inversion bypasses the regulated fructose-1,6-bisphosphate intermediate and the inhibitory fructose bisphosphatase step, achieving net fructose-6-phosphate production despite high glucose/sucrose conditions.

Inventive Principle:

Principle #13The other way round (Inversion)

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

Enables the production of fructose-6-phosphate even at high glucose or sucrose concentrations by utilizing unregulated enzymes, enhancing the efficiency of the conversion process.

Implementation Method 1

enzymatically converting dihydroxyacetone phosphate (DHAP) into dihydroxyacetone (DHA)

Methodology Applied

Scientific EffectEnzymatic conversion: Enzyme

Implementation Method 2

enzymatically converting the thus produced dihydroxyacetone (DHA) together with glyceraldehyde-3-phosphate (G3P) into fructose-6-phosphate (F6P)

Methodology Applied

Scientific EffectCondensation reaction: Chemical Bonding

Data Source

PatentUS11773421B2Method for producing fructose-6-phosphate from dihydroxy acetone phosphate and glyceraldehyde-3-phosphate

Publication Date: 2023.10.03 GLOBAL BIOENERGIES

AI summary

Described is a method for the production of fructose-6-phosphate (F6P) from dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (G3P) comprising the steps of:(a) enzymatically converting dihydroxyacetone phosphate (DHAP) into dihydroxyacetone (DHA); and(b) enzymatically converting the thus produced dihydroxyacetone (DHA) and glyceraldehyde-3-phosphate (G3P) into fructose-6-phosphate (F6P); orcomprising the steps of:(a′) enzymatically converting glyceraldehyde-3-phosphate (G3P) into glyceraldehyde; and(b′) enzymatically converting the thus produced glyceraldehyde together with dihydroxyacetone phosphate (DHAP) into fructose-1-phosphate (F1P); and(c′) enzymatically converting the thus produced fructose-1-phosphate (F1P) into fructose-6-phosphate (F6P).