Genetically Modified Pseudomonas putida S12 for HMF-Acid Bioconversion

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Solution Overview

Problem

The accumulation of 5-(hydroxymethyl)furan-2-carboxylic acid (HMF-acid) is a challenge in the bioproduction of furan-2,5-dicarboxylic acid (FDCA) and biofuel production from lignocellulose, as it requires long process times or alternative measures for removal, which is undesirable for many applications.

Innovation Solution

Expression of specific polypeptides, such as those with amino acid sequences found in SEQ ID NO: 1, 2, 3, or 4, and their analogues, in genetically modified cells like Pseudomonas putida S12, enhances the bioconversion of HMF-acid, reducing its accumulation and improving the biocatalytic transformation of furanic compounds.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If the hmfH gene from Cupriavidus basilensis HMF14 is functionally introduced in Pseudomonas putida S12, then FDCA production capability is improved, but HMF-acid accumulates requiring long process times or alternative removal measures

Engineering Contradiction:

ImproveFDCA production capabilityVSAvoidprocess time

Core Design Contradiction:

ProductivityVSLoss of time

Solution Approach 1:

The invention segments the HMF-acid management problem by introducing a separate functional component (HMF-acid removing polypeptide) that specifically addresses HMF-acid removal, allowing the main hmfH gene to focus on FDCA production. This division of functional responsibilities enables simultaneous optimization of both FDCA production and HMF-acid removal without requiring long process times.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The HMF-acid removing polypeptide acts as an intermediary that handles the problematic HMF-acid intermediate, preventing its accumulation and enabling the main biocatalytic pathway to proceed efficiently toward FDCA production. This intermediary component resolves the bottleneck created by HMF-acid buildup.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Productivity

Engineering Contradiction:

ImproveFDCA production capabilityVSAvoidHMF-acid accumulation

Core Design Contradiction:

ProductivityVSObject-generated harmful factors

Solution Approach 1:

The invention converts the harmful accumulation of HMF-acid into a beneficial process by introducing a polypeptide that actively removes HMF-acid. The harmful intermediate is transformed from a process bottleneck into a managed substrate that is continuously converted to desired products, turning a disadvantage into an advantage.

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

Solution Approach 2:

The HMF-acid removing polypeptide serves as an intermediary that specifically targets and removes the harmful HMF-acid intermediate, preventing its accumulation while allowing the main biocatalytic pathway to continue producing FDCA. This intermediary function resolves the harmful effect without disrupting the beneficial FDCA production.

Inventive Principle:

Principle #24Intermediary (Mediator)

3Productivity

If specific polypeptides are expressed in genetically modified cells, then HMF-acid bioconversion is enhanced, but device complexity increases

Engineering Contradiction:

ImproveHMF-acid bioconversion efficiencyVSAvoidgenetic modification complexity

Core Design Contradiction:

ProductivityVSDevice complexity

Solution Approach 1:

The HMF-acid removing polypeptide is designed to perform a specific universal function (HMF-acid removal) that can be applied across different biocatalytic systems. By creating a modular, specialized component with a single clear function, the invention achieves enhanced bioconversion efficiency while keeping the added complexity manageable and well-defined.

Inventive Principle:

Principle #6Universality (Multi-functionality)

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

This approach effectively minimizes HMF-acid accumulation, allowing for increased FDCA production in shorter process times and improved bioproduction of chemicals, enhancing the efficiency of biofuel and biochemical production from lignocellulosic feedstocks.

Implementation Method 1

The first polypeptide is capable of transporting 5-(hydroxymethyl)furan-2-carboxylic acid (HMF-acid)

Methodology Applied

Scientific EffectActive transport:

Implementation Method 2

a second polynucleotide sequence coding for a second polypeptide having HMF-acid converting activity

Methodology Applied

Scientific EffectEnzymatic bioconversion: Enzyme

Data Source

PatentUS10023890B2Genetically modified cell and process for use of said cell

Publication Date: 2018.07.17 PURAC BIOCHEM BV

AI summary

The present invention relates to the field of biotransformation of furanic compounds. More particular the present invention relates to novel genetically modified cells with improved characteristics for biocatalytic transformation of furanic compounds and a vector suitable for the genetic modification of a host cell. Further aspects of the invention are aimed at processes for biotransformation of 5-(hydroxymethyl)furan-2-carboxylic acid (HMF-acid) and its precursors with the use of the cell according to the invention.