Pyrrolidone carboxyl peptidase, commonly known as PYRase, is an exopeptidase that catalytically cleaves an N-terminal pyroglutamic acid from peptides or proteins. The diverse functions of PYRases in bacterial enzymology have prompted the development of various bacterial diagnostic techniques. However, the specific physiological role and activity of this enzyme across the bacterial kingdom remain unclear. Here, we present a functional phenoxy-1,2-dioxetane chemiluminescent probe (PyrCL) that can selectively detect PYRase activity in both Gram-positive and Gram-negative bacteria. The probe activation mechanism is based on the cleavage of a pyroglutamyl substrate, followed by a release of the phenoxy-dioxetane luminophore, which then undergoes efficient chemiexcitation to emit a green photon. Probe PyrCL exhibits an effective turn-on response with superior detection capability in terms of response time and sensitivity compared to existing fluorescence probes. The superior detection sensitivity of the chemiluminescent probe enables us to reveal previously undetected PYRase activity in Streptococcus mutans. Furthermore, it enables the discrimination of Pseudomonas aeruginosa from other Gram-negative bacteria in the tested panel, based on their distinct PYRase activity. We expect that probe PyrCL will have great value for PYRase-based bacteria diagnosis with use in basic research and clinical applications.

中文翻译：

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高响应化学发光探针揭示铜绿假单胞菌中独特的 PYRase 活性

吡咯烷酮羧基肽酶，通常称为 PYRase，是一种肽链外切酶，可催化从肽或蛋白质中裂解N末端焦谷氨酸。 PYRases 在细菌酶学中的多种功能促进了各种细菌诊断技术的发展。然而，这种酶在细菌界的具体生理作用和活性仍不清楚。在这里，我们提出了一种功能性苯氧基-1,2-二氧杂环丁烷化学发光探针（PyrCL），它可以选择性检测革兰氏阳性和革兰氏阴性细菌中的 PYRase 活性。探针激活机制基于焦谷氨酰底物的裂解，随后释放苯氧基二氧杂环丁烷发光体，然后该发光体经过有效的化学激发以发射绿色光子。与现有荧光探针相比，PyrCL 探针表现出有效的开启响应，在响应时间和灵敏度方面具有卓越的检测能力。化学发光探针卓越的检测灵敏度使我们能够揭示变形链球菌中以前未检测到的 PYRase 活性。此外，它还可以根据铜绿假单胞菌独特的 PYRase 活性，将其与测试组中的其他革兰氏阴性细菌区分开来。我们预计探针 PyrCL 对于基于 PYRase 的细菌诊断在基础研究和临床应用中具有巨大的价值。