Hydrophobins are small surface-active amphiphilic proteins that contain both hydrophobic and hydrophilic areas on their surface. They can self-assemble to form a stable film at the interface of both polar and non-polar solution phases. Thanks to high surface activity and amphiphilicity, Hydrophobins have broad application prospects as surfactants in interface engineering, emulsifiers in food processing, bioimmobilization agents in nanotechnology, and powerful protein purification tags in biotechnology. In summary, hydrophobins are vital tools for today's bioengineering and synthetic biology.
Our project features BslA, a recently discovered bacterial-origin hydrophobin with great potential and wide application in synthetic biology. Retaining all the virtues as a hydrophobin, BslA is much easier to produce compared to their fungal-origin cousins, which overcomes the limitations of their industrial use. We hope to unleash the potential of BslA and solve some of the major concerns of the synbio community and humanity as a whole. To this end, Our team designed three BslA fusion proteins that target three problems: protein purification, antibiotic abuse, and plastic pollution.
The first problem we aim to solve is related to protein purification, an essential technique in synthetic biology. An emerging approach to protein separation and purification is the Aqueous two-phase separation (ATPS). ATPS involves building a system of two liquid phases. The target protein fuses with a hydrophobin tag, which changes the hydrophobicity of the protein. The fusion protein will thus simultaneously move to the interface of the two liquid phases. However, ATPS has yet to be adopted in industrial production because of two drawbacks: the high costs of fungal hydrophobin and the low separation speed (~10 hours/batch). If we can solve ATPS's dependence on fungal hydrophobin and implement a continuous flow system, ATPS will become a fast and highly efficient method to purify proteins.
The second problem is antibiotic abuse. Over the past decade, the world has witnessed an alarming increase in antibiotic usage. Antibiotic abuse leads to increased bacterial resistance. New super-resilient bacteria impel scientists to develop new antibiotics, which inflicts enormous R&D costs.
The third problem is plastic pollution. Plastic pollution has become a critical threat to the environment and human health. Polyethylene terephthalate (PET) is one of the most commonly used plastics and a major source of plastic pollution. Currently, we use physical, chemical, and biological approaches to recycle PET. However, physical and chemical recycling has many limitations, such as the inability to achieve closed-loop recovery and a high chance of secondary pollution.
Our team is trying to improve traditional ATPS by incorporating a continuous-flow system and replacing fungal hydrophobins with BslA. We will construct BslA fusion proteins and purify the target proteins using the improved ATPS method. We are confident in achieving a yield higher than traditional ATPS at a faster speed and lower cost.
Scientists have proposed different solutions to antibiotic abuse, such as combination drugs, quinolones, and antimicrobial peptides. Antimicrobial peptides have broader application prospects because of their exceptional biocompatibility, low chance to render bacteria drug resistance, and fast sterilization. Our research focuses on LL-37, an antimicrobial peptide from the Cathelicidin family found in human cells. Studies have shown that hydrophobins help increase the binding ability of antimicrobial peptides and thus help them eliminate bacteria more efficiently. With the LL37-BslA fusion protein, we are striving to enhance the power of this antimicrobial peptide and counter antibiotic abuse.
As discussed in the problem, Physical and chemical solutions to plastic pollution have many limitations (see the previous section). In contrast, biological recycling is a promising method for its high efficiency, closed-loop recycling, and low secondary pollution. Biodegrading PET consists of two processes: adsorption and degradation. During adsorption, the enzyme mPETase binds to the PET film. However, the surface of mPETase is hydrophilic while the PET film is hydrophobic, so the low affinity between them reduces the adsorption efficiency. After intensive literature research, we are inspired to enhance adsorption rate and degradation efficiency by using hydrophobins to increase binding efficiency. We have constructed the mPETase-BslA fusion protein to explore and expand the mPETase's potential in PET degradation.
As a rising star in the hydrophobin family, BslA has unlimited potential. Our team aims to harness BslA's power and solve the three problems in the synbio community and humanity. We hope our project can reduce the cost of protein purification, help improve the efficiency of LL37 as an antimicrobial peptide, help improve the efficiency of mPETase and reduce the environmental pollution caused by plastic. At the same time, we are also exploring more features belonging to BslA during the experiment to deepen our understanding of it.
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