A competitive high-throughput screening platform for designing polylactic acid-specific binding peptides
Lu, Y., Hintzen, K. W., Kurkina, T., Ji, Y., Schwaneberg, U., Advanced Science. https://doi.org/10.1002/advs.202303195.
A 96-well microtiter plate based high-throughput screening system for polylactic acid (PLA) specific binding was developed for detection, sorting, and material-specific degradation of PLA in mixed plastics.
Polylactic acid (PLA) is a rapidly-growing bioplastic with broad applications in food packaging, agricultural films, and drug delivery. To meet the application requirements, PLA is often blended with polypropylene (PP) to enhance its melt processability, thermal stability, and stiffness. The development of plastic recycling processes in the future will be essential to separate PLA from PP in mixed plastics. Material binding peptides (MBPs) have outstanding binding properties to a variety of materials. Engineered MBPs that can bind to polymers in a material-specific manner have a high potential for material-specific detection or enhanced degradation of PLA in mixed PLA/PP plastics. To obtain a material-specific MBP for PLA binding (termed PLAbodies), protein engineering of MBP Cg-Def for improved PLA binding specificity is reported in this work. A 96-well microtiter plate based high-throughput screening system for PLA specific binding (PLABS) was first developed and validated in a protein engineering (KnowVolution) campaign. As a result, a Cg-Def variant V2 (Cg-Def S19K/K10L/N13H) with a 2.3-fold improved PLA binding specificity compared to PP was obtained. Contact angle and surface plasmon resonance measurements confirmed improved material-specific binding of V2 to PLA (1.30-fold improved PLA surface coverage). The established PLABS screening platform for PLA bodies designing can be applied in detection, sorting, and material-specific degradation of PLA in mixed plastics.
Yi Lu is financially supported by the China Scholarship Council (CSC) scholarship. This work was supported by European Union's Horizon 2020 research and innovation program under grant agreement no. 870294 for the project MIX-UP.
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Lu, Y., Hintzen, K. W., Kurkina, T., Ji, Y., Schwaneberg, U., Advanced Science. https://doi.org/10.1002/advs.202303195.
Enhanced PLA-specific binding of Cg-Def by KnowVolution campaign comprising four phases. In Phase I, potentially beneficial positions were identified by screening cepPCR library of Cg-Def using a 96-well microtiter plate based high-throughput screening system. Then in Phase II, beneficial positions were determined after screening the SSM libraries of identified positions. In Phase III, the computational analysis of determined beneficial positions and substitutions was performed to find compatible substitutions. Finally, a recombined variant was generated and characterized for PLA binding specificity by competitive binding tests in one pot, contact angle, and SPR measurements in Phase IV. PLA: polylactic acid; MBP: material binding peptide; PP: polypropylene; cepPCR: casting error-prone polymerase chain reaction method; SPR: surface plasmon resonance; SSM: site-saturation mutagenesis