Hydrophobins are small surface-active proteins, and have both fungal and bacterial origins. Hydrophobins originated from fungi are divided in to two class, and are being widely used and applied.
BslA is the only bacterial hydrophobin identified so far. It is similar in properties to fungal hydrophobins.
Hydrophobin’ s hydrophobic part is exposed on the surface forming a planar area called the hydrophobic patch, making the surface of the protein contain both hydrophobic and hydrophilic area therefore making the surface making the hydrophobins amphiphilic. Because of the hydrophobins’ amphiphilicity property it can self-assemble themselves with others.
In 2022, the TJUSLS_China discovered a mutant super5, which is mPETase (BBa_K3715005). The degradation efficiency of mPETase is 163 times that of wild type, which greatly improves the degradation efficiency of PET. Therefore, this year we decided to further improve mPETase with excellent degradation efficiency.
Our team is devoted to increasing PET degradation efficiency and improving mPETase. Therefore, we proposed the construction of a fusion protein of mPETase and BslA, hoping to improve the PET degradation efficiency by improving the adsorption capacity of mPETase enzyme on the hydrophobic PET film.
By constructing the mPETase-GSlinker-BslA and mPETase, the PET degradation efficiency will be enhanced due to the unique properties of amphiphilicity and self-assembly of hydrophobins.
To sum up the above, we use our practice to make some contribution to PET degradation in order to protect the environment.
For molecular cloning, we selected pET28a as vector. We successfully amplified two gene segments of mPETase (as control group), mPETase-GSlinker-BslA (Figure 1a). Then we digested and connected all the segments to pET28a vector through two restriction enzymes of BamHI and XhoI. At present, two recombinant plasmids have been successfully constructed (Figure 1b).
[1] Puspitasari, Nathania, Shen-Long Tsai, and Cheng-Kang Lee. "Fungal hydrophobin RolA enhanced PETase hydrolysis of polyethylene terephthalate." Applied Biochemistry and Biotechnology 193.5 (2022): 1284-1295.
[2] Ribitsch, Doris, et al. "Enhanced cutinase-catalyzed hydrolysis of polyethylene terephthalate by covalent fusion to hydrophobins." Applied and Environmental Microbiology 81.11 (2015): 3586-3592.
