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Filamentous fungi detecting and disarming furfural


Furfural is a small furan compound commonly found in lignocellulosic waste. Furfural greatly inhibits the growth of microorganisms, thus the presence of furfural makes it extremely cumbersome to utilize lignocellulosic waste as a renewable carbon source for production in biomanufacturing.

DTU BioBuilders are engineering a filamentous fungus to detect and convert furfural into harmless compounds. To detect furfural, we constructed a Synthetic Expression System (SES) containing a synthetic promoter and a synthetic transcription factor (sTF). In addition, we identified a promoter native to A. niger that is highly inducible in response to furfural and can serve as a furfural biosensor. With our engineered fungus, we aim to unlock the potential of lignocellulosic waste as a cheap and renewable carbon source in fermentation, thereby promoting sustainable production and responsible use of our shared resources on Earth.

The chemical structure of furfural and its GHS hazard classifications

The Problem

Depletion of resources and growing piles of waste


The human population is growing. The food demand increases while we are running out of space and resources. At the same time, the piles of lignocellulosic waste from agriculture and forestry are growing and managed unsustainably, causing harm to human health and the environment. We need to use our resources more efficiently and recycle any waste we generate in the most efficient way to combat these issues.

Lignocellulosic waste has a huge potential as a renewable and sustainable carbon resource in microbial fermentation for production of compounds with high value. However, lignocellulosic waste cannot currently be efficiently utilized for this purpose, due to the presence of a highly toxic compound: furfural.

Our Solution

Aspergillus niger detecting and converting furfural




DTU BioBuilders aim to combat furfural in lignocellulosic waste using engineered Aspergillus niger capable of detecting and converting furfural into harmless compounds. Our method to do the same is showcased in the flowchart below.

Developing a biosensor for detecting furfural


To our knowledge, no biosensor capable of detecting furfural exists. Therefore, we needed to develop our own. We used two different strategies:

  • Rationally modifying a synthetic transcription factor proven to sense furfural-like compounds in yeast.
  • Identifying and using native promoters within A. niger that we predicted would be activated by furfural.

The pipeline to developing these two processes are shown below.

AlphaFold2 model of sBAD structure (Interactive)

Detecting furfural by modifying synthetic transcription factors (sTFs)

As a starting point, we constructed sTFs with a corresponding synthetic promoter using the benzoic acid-sensing sTF, sBAD, which has been developed in yeast. Since the structure of benzoic acid resembles that of furfural, we anticipated sBAD could be used as a furfural-sensing biosensor. We introduced mutations in the sensing domain of sBAD, that we predicted would improve the binding affinity to furfural using Rosetta software. This system is one of very few reported SESs for filamentous fungi and the first in iGEM history!

Detecting furfural using a native promoter

We identified a gene in A. niger that is highly upregulated in response to furfural. We successfully constructed a plasmid expressing mCherry from the promoter of this upregulated gene and results indicate that mCherry expressed from the plasmid was also upregulated in response to furfural. Nonetheless, further investigations are needed to confirm that this is the first fully functional furfural biosensor.

Converting furfural into harmless compounds



We introduced the heterologous genes Arz_7774 and hmfH into A. niger. These enzymes have been reported to convert furfural into the relatively harmless compound furoic acid. The gene Arz_7774 is a putative NAD-dependent aldehyde dehydrogenase from the remarkable kerosene fungus Amorphotheca resinae ZN1 known as the jet fuel fungus. We also tested the gene hmfH, a putative FAD-dependent oxidoreductase from the soil bacterium Cupriavidus basilensis HMF14.

However... Real solutions go beyond the lab!


If solutions don't go beyond the doors of the lab, they are not real solutions. Therefore, iGEM is so much more than the science itself! DTU BioBuilders did excessive work in outreach and education, human practices to ensure that our project embraced the needs of the industry thereby conveying as much benefit to the world as possible.
Furthermore, DTU BioBuilders acknowledge that scientists need to collaborate in order to reach impactful solutions to the many global challenges that the world faces and those yet to come. For that reason we initiated multiple collaborations, and even formed a partnership with TU-Delft to benefit from each other's strengths. Our project is highly centered around sustainability, and we explicitly try to address the SDGs.

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