Microbial Host Cell Adaptation for Terpene Tolerance

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Solution Overview

Problem

Biotechnological production of terpenoids is hindered by the toxicity of terpenoids towards microorganisms, leading to low product titers and growth arrest, necessitating the development of host cells with increased tolerance to these substances.

Innovation Solution

Adaptation of E. coli cells through constant growth in the presence of toxic terpenes like Isoprenol, followed by selection pressure to generate mutations that increase tolerance, which are then introduced into wild-type and knockout cells to reduce toxicity and enhance production potential.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Quantity of substance

If terpenoids are produced in microorganisms, then product titer increases, but toxicity towards microorganisms increases leading to growth arrest

Engineering Contradiction:

Improveproduct titerVSAvoidtoxicity towards microorganisms

Core Design Contradiction:

Quantity of substanceVSObject-affected harmful factors

Solution Approach 1:

The patent applies evolution engineering to convert the harmful effect of terpene toxicity into a beneficial adaptation. By subjecting microorganisms to progressive terpene stress over multiple generations, the system selects for mutants with increased tolerance, ultimately converting the toxic substance into a manageable production condition that enables higher titers

Inventive Principle:

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

Solution Approach 2:

The patent changes the physiological parameters of the microorganism through evolutionary adaptation. By exposing cells to increasing terpene concentrations over generations, the system induces genetic mutations that alter membrane composition and other physiological parameters, resulting in strains with fundamentally changed tolerance thresholds

Inventive Principle:

Principle #35Parameter changes

2Productivity

If host cells are engineered to produce terpenoids, then production potential increases, but tolerance to terpenes decreases

Engineering Contradiction:

Improveproduction potentialVSAvoidtolerance to terpenes

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The patent applies preliminary action by pre-adapting host cells to terpene stress through evolution engineering before actual production. This preliminary exposure to sub-lethal terpene concentrations over multiple generations prepares the cells with enhanced tolerance mechanisms, ensuring they can withstand the full production load without growth arrest

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent implements feedback through iterative cycles of terpene exposure and selection. Each generation of cells is exposed to terpene stress, the survivors are selected, and this information feeds into the next generation's adaptation, creating a progressive feedback loop that continuously enhances tolerance while maintaining production capability

Inventive Principle:

Principle #23Feedback

Data Source

PatentUS20230041211A1Decreasing toxicity of terpenes and increasing the production potential in micro-organisms

Publication Date: 2023.02.09 BASF SE

AI summary

The invention pertains to novel methods to increase the tolerance of microbial host cells to toxic substance, for example terpenes and alcohols and other membrane disrupting substances, as well as modified organism with such an increased tolerance a compared to the unmodified organism.