Dihydropyrimidinase from Saccharomyces kluyveri can hydrolyse polyamides

• Published in: Frontiers in bioengineering and biotechnology Vol. 11; p. 1158226
• Main Authors: Quartinello, Felice, Subagia, Raditya, Zitzenbacher, Sabine, Reich, Johanna, Vielnascher, Robert, Becher, Erik, Hall, Mélanie, Ribitsch, Doris, Guebitz, Georg M
• Format: Journal Article
• Language: English
• Published: Switzerland Frontiers Media S.A 26.04.2023
• Subjects: Bioengineering and Biotechnology; dihydropyrimidinase; enzyme; hydrolysis; polyamide; Saccharomyces kluyveri; surface functionalization; enzyme; polyamide; dihydropyrimidinase; Saccharomyces kluyveri; surface functionalization; hydrolysis
• ISSN: 2296-4185; 2296-4185
• DOI: 10.3389/fbioe.2023.1158226

In dihydropyrimidinase (DHPaseSK) is involved in the pyrimidine degradation pathway, which includes the reversible ring cleavage between nitrogen 3 and carbon 4 of 5,6-dihydrouracil. In this study, DPHaseSK was successfully cloned and expressed in BL-21 Gold (DE3) with and without affinity tags. Thereby, the Strep-tag enabled fastest purification and highest specific activity (9.5 ± 0.5 U/mg). The biochemically characterized DHPaseSK_Strep had similar kinetic parameters (K /K ) on 5,6-dihydrouracil (DHU) and -nitroacetanilide respectively, with 7,229 and 4060 M  s . The hydrolytic ability of DHPaseSK_Strep to polyamides (PA) was tested on PA consisting of monomers with different chain length (PA-6, PA-6,6, PA-4,6, PA-4,10 and PA-12). According to LC-MS/TOF analysis, DHPaseSK_Strep showed a preference for films containing the shorter chain monomers (e.g., PA-4,6). In contrast, an amidase from (NFpolyA) showed some preference for PA consisting of longer chain monomers. In conclusion, in this work DHPaseSK_Strep was demonstrated to be able to cleave amide bonds in synthetic polymers, which can be an important basis for development of functionalization and recycling processes for polyamide containing materials.