Description
Problem
The ongoing climate crisis is mainly driven by the lack of an efficient renewable energy technology, which would serve as a substitute to the excessive use of non-renewable fossil fuels. Combustion of fossil fuels such as coal, oil and natural gas releases carbon dioxide and other greenhouse-gases into the atmosphere causing global climate change and air pollution [1]. While currently available solar and wind energy technologies are developing rapidly, the modern renewable energy sources are highly dependable on environmental conditions and thus have considerable geographical and climate limitations.
This instability underscores the critical need for efficient energy storage solutions to ensure a stable and reliable power supply. Lithium-ion batteries have emerged as a leading technology in this context. They play a key role in people's everyday life: they power our phones, laptops, smartwatches, vacuum cleaners, cars, and more. Despite their widespread use and our growing dependence on them, the production and use of Li-ion batteries have significant environmental impacts. In particular, lithium mining can cause severe damage to ecosystems and contribute to freshwater pollution — threatening one of humanity's most essential resources [2].
Constant availability of clean freshwater is critical for human wellbeing and survival. But almost a quarter of the world's population does not have continuous access to clean drinkable water, which causes over half a million deaths a year [3]. Taking into account the constantly increasing demand for the freshwater supply, wastewater management paradigm is getting shifted from “treatment and disposal” to “reuse, recycle and resource recovery”. However, wastewater treatment requires a lot of resources and consumes a lot of electricity. Up to 5% of the world's produced electricity is used for wastewater treatment [4].
However, this also presents an opportunity to address both challenges by developing an eco-friendly battery that can simultaneously treat wastewater, as various organic compounds in wastewater can be utilized to generate electricity. The systems that can do this are called microbial fuel cells, MFCs, or simply biobatteries. These batteries are powered not by Li-ions, but by microorganisms that consume organic matter from wastewater and produce electrons. These electrons are then transferred to electrodes, generating electrical current. In the end, we get “free electricity”, treated water, and saved resources, making wastewater facilities more affordable in the underdeveloped regions [3]. Integrating innovative technologies like MFCs into wastewater treatment contributes not only to improving water quality and sustainable resource use but also to achieving global targets for clean energy access and environmental resilience.
In a typical MFC setup (Figure 1), anode chamber is filled with living cells, which consumes organic substrate and transfers electrons to anode. Electrons then move from the anode through an external circuit to the cathode, where they participate in the reduction of a terminal electron acceptor, such as potassium hexacyanoferrate. A proton (cation) exchange membrane separates the two chambers, enabling proton migration while preventing the reverse diffusion of oxygen or electron acceptors into the anode, thereby maintaining stable redox conditions and continuous current generation.
Figure 1. Typical MFC setup. The anode chamber contains microorganisms that metabolize organic compounds and release electrons and protons. Electrons are transferred to the anode either directly or with the aid of electron transfer mediators (e.g., methylene blue). The anode and cathode chambers are separated by a proton exchange membrane (PEM), which allows protons to migrate from the anode to the cathode chamber. Meanwhile, electrons flow through an external circuit to the cathode, where they participate in the reduction of a terminal electron acceptor (e.g., potassium hexacyanoferrate).
However, many limitations exist in operating MFCs. Most MFCs have low power density and efficiency, can operate only for a limited time—partly due to fluctuations in substrate supply and composition—face challenges in scaling up electrode size, and are sensitive to changes in environmental conditions [5].
Our idea
To enhance the viability of the fuel cells, we selected the widely used yeast Saccharomyces cerevisiae . While bacteria are often the preferred choice for constructing MFCs due to their rapid growth and inherent extracellular electron transfer capabilities, S. cerevisiae offers several significant advantages. Firstly, yeast is a well-established eukaryotic model organism with a long history of use in molecular biology and biotechnology, meaning its genetics and physiology are exceptionally well understood. This facilitates experimental manipulation, genetic engineering, and the interpretation of results. Secondly, S. cerevisiae possesses a robust cell wall that enhances its overall resilience. In addition, yeast can grow under both aerobic and anaerobic conditions (the anode chamber is typically anaerobic) and can tolerate a wide range of pH values as well as high ethanol concentrations. Altogether, these characteristics contribute to enhanced viability and stability during long-term operation compared to most bacterial species [6] [7].
Our project focuses on engineering yeast S. cerevisiae, capable of electron transfer towards electrodes, to generate eclectic current. In wild-type Saccharomyces cerevisiae, much of the reducing power is consumed through fermentation pathways—producing ethanol and glycerol—or directed into the mitochondrial respiratory chain, rather than being transferred to the electrode. The goal of yeast engineering is to increase nicotinamide adenine dinucleotide (NADH) pool in the cytosol, which serves as the main electron carrier for electricity generation in microbial fuel cells. Our strategy aims to modify S. cerevisiae to serve as a more efficient biocatalyst for microbial fuel cells (MFCs).
To achieve these goals, our project involves modifying yeast central metabolism to enhance cell capacity for electron transfer to anode by several ways.
Deletion of NADH-consuming genes with CRISPR-Cas9 (Figure 2):
- Glycerol-3-phosphate dehydrogenase (GPD1/2): generates glycerol [8];
- Alcohol dehydrogenase (ADH1/2/5): converts acetaldehyde into ethanol [9];
- NADH dehydrogenase (NDE1/2): catalyzes the oxidation of cytosolic NADH, providing cytosolic NADH to the mitochondrial respiratory chain [10];
Figure 2. Modifications in the yeast central metabolism. GPD1, GPD2, ADH1,5 and NDE1,2 genes were deleted to increase the cytosolic pool of NADH
We propose that the increase in the cytosolic NADH pool can be efficiently utilized, with the aid of electron transfer mediators such as methylene blue shuttling electrons from intracellular NADH to the electrode surface, to power an electrical circuit.
Expression of codon-optimized fungal cellobiose dehydrogenase (CDH).
To enhance the yeast's capacity for wastewater treatment and to further facilitate electron transfer to the anode, we plan to integrate into the yeast genome a cellobiose dehydrogenase (CDH) gene derived from the fungus Phanerochaete chrysosporium. CDH overexpression broadens the range of substrates that the cell can metabolize. In the context of microbial fuel cells, expressing CDH enables S. cerevisiae to catalyze the oxidation of various organic compounds while directly transferring electrons to the electrode, either through direct electron transfer or via redox mediators. This should significantly improve the efficiency of bioelectricity generation and enhance the degradation of organic pollutants in wastewater [11]. In addition, yeast cells displaying fully functional CDH on their surface may eliminate the need for electron transfer mediators such as methylene blue, or at least significantly reduce their required amount, making the entire system more eco-friendly.
Together, the genetic modifications that we introduced into yeast cells have a purpose to redirect electron flow away from ethanol and glycerol production towards extracellular electron transfer, thus boosting the generation of electrical current.
Figure 3. Proposed MFC design. Increasing the intracellular NADH pool together with the overexpression of cellobiose dehydrogenase (CDH) on the yeast cell surface is expected to enhance the transfer of electrons from yeast cells to the anode.
Microbial fuel cell design has relied on trial-and-error strain screening with no predictive framework connecting genetics to power output. We introduce the mechanistic multi-scale model that bridges this gap: cellular NADH flux models predict electron surplus from gene deletions, biofilm kinetics translate single-cell metabolism into collective current dynamics, and reactor optimization maximizes power across biological and electrochemical timescales.
Following successful genetic modifications, the engineered strains were integrated into custom-built MFC hardware, allowing measurement of the battery's performance and electrical characteristics. We designed our MFC using acrylic chambers so the system is easy to build, test and visualise. We used carbon felt electrodes that give microbe space to grow inside anode chamber. The design is compact, reduces leakage, and can be flexibly adjusted for future experiments.
Utilising this system allows us to improve the efficiency of yeast-based microbial fuel cells and provides the way for renewable, bio-based electricity production.
Final product
The final product of the project is a functioning microbial fuel cell which consists of genetically modified yeast strains with optimized metabolic pathways integrated into biocompatible battery hardware.
The modified yeast strains possess an increased cytosolic NADH pool, achieved through rational design and targeted genetic engineering. Electrons from intracellular NADH can be efficiently transferred to the electrodes via the CDH enzyme displayed on the yeast cell surface, which acts as a direct link between cellular metabolism and electricity generation. An ideal microbial fuel cell (MFC) would not only generate power efficiently but also function as an effective system for wastewater treatment.
