Introduction
The accumulation of plastics, among others PET, poses a serious threat to the environment due to its extensive and short-term use and insufficient recycling processes. Microbial degradation of PET can contribute to combat this problem. One promising organism is Comamonas thiooxydans S23, since it is capable to import and metabolise TPA, a degradation product of PET. Also, a reporter strain of C. thiooxydans S23 namely C. thiooxydans ReporTPA_UHH04 has already been constructed by Dierkes et al. (2022), which exhibits TPA induced fluorescence of sfGFP. [1]
Goal
We aim to equip C. thiooxydans ReporTPA_UHH04 with suitable PETases to enable it to degrade PET to TPA. To optimize expression and secretion, different promotors and signal peptides will be compared. The strains will be grown as biofilms on PET-film as well as with PET-powder and should be able to directly report PET degradation through fluorescence. This should serve as a first proof of concept of C. thiooxydans being able to degrade PET to TPA, to further modify the strain to finally being able to convert TPA into a useful platform chemical in later work.
Methods
The formation of biofilms was visualised via confocal laser scanning microscopy and scanning electron microscopy. The performance of the PET degrading reporter strains for degrading PET as well as the TPA induced fluorescence itself will be analysed through confocal laser scanning microscopy and microtiter plate assays. PET degradation products will furthermore be verified through HPLC and MS.
Results
We were already able to identify conditions under which the C. thiooxydans could form biofilms on PET film. Further, potential PETases were already cloned into ReporTPA_UHH04 and right now their general activity is getting evaluated. The degradation ability of the promising strains to degrade PET as a biofilm on PET film as well as for PET powder will be examined in upcoming experiments.
Conclusion
C. thiooxydans can form biofilms on PET and if it can successfully be employed to degrade PET in different forms to TPA, the strain is well suited to be further manipulated to convert the imported through PET degradation released TPA into useful platform chemicals.
[1] Dierkes RF et al., Appl Environ Microbiol. 2023 Jan 31;89(1):e0160322. doi: 10.1128/aem.01603-22.