In plasma-driven biocatalysis, enzymes are employed to carry out reactions using species generated by non-thermal plasmas as the precursors. We have previously demonstrated that this is feasible in principle, but that the approach suffers from the short lifetime of the biocatalyst under operating conditions. In this work, protection strategies were investigated to prevent the dielectric barrier discharge plasma-induced inactivation of biocatalysts, using recombinant unspecific peroxygenase from Agrocybe aegerita( rAaeUPO), one of the most promising enzymes for plasma-driven biocatalysis. Treatment in oxygen-free atmospheres did not provide any advantage over treatment in synthetic air, indicating that the detrimental reactive species did not originate from oxygen in the plasma phase. Chemical scavengers were employed to eliminate undesired reactive species, without any long-term effect on enzyme lifetime. Similarly, chaperones, including the known stress response proteins Hsp33, CnoX, and RidA did not increase the lifetime of rAaeUPO. Immobilization of the biocatalyst proved effective in preserving enzyme activity. The residual activity of rAaeUPO after plasma treatment strongly depended on the specific immobilization support. Essentially complete protection for at least 15 min of plasma exposure was achieved with an epoxy-butyl-functionalized carrier. This study presents new insights into plasma–protein interactions and plots a path forward for protecting biocatalytic proteins from plasma-mediated inactivation.

在等离子体驱动的生物催化中，使用酶来进行反应，该反应使用由非热等离子体产生的物质作为前体。先前我们已经证明了这在原则上是可行的，但是该方法在操作条件下遭受生物催化剂寿命短的困扰。在这项工作中，研究了使用Agrocybe aegerita（r Aae）的重组非特异性过氧化酶来防止介电势垒放电等离子体诱导的生物催化剂失活的保护策略。UPO），是血浆驱动生物催化最有前途的酶之一。与在合成空气中进行处理相比，在无氧气氛中进行处理没有提供任何优势，这表明有害的反应性物种并非源自等离子体相中的氧气。使用化学清除剂消除不需要的反应性物质，而对酶寿命没有任何长期影响。同样，伴侣蛋白，包括已知的应激反应蛋白Hsp33，CnoX和RidA，也不会延长r Aae UPO的寿命。证明固定化生物催化剂对保持酶活性有效。r Aae的残留活性血浆处理后的UPO强烈依赖于特定的固定化支持。使用环氧丁基官能化的载体可实现对至少15分钟的等离子体暴露的基本完全保护。这项研究提供了对血浆蛋白相互作用的新见解，并为保护生物催化蛋白免受血浆介导的失活提供了一条途径。