Pak Valeriy V, Ezeriņa Daria, Lyublinskaya Olga G, Pedre Brandán, Tyurin-Kuzmin Pyotr A, Mishina Natalie M, Thauvin Marion, Young David, Wahni Khadija, Martínez Gache Santiago Agustín, Demidovich Alexandra D, Ermakova Yulia G, Maslova Yulia D, Shokhina Arina G, Eroglu Emrah, Bilan Dmitry S, Bogeski Ivan, Michel Thomas, Vriz Sophie, Messens Joris, Belousov Vsevolod V
Department of Metabolism and Redox Biology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997, Russia; Institute for Cardiovascular Physiology, Georg August University Göttingen, Göttingen 37073, Germany.
VIB-VUB Center for Structural Biology, Vlaams Instituut voor Biotechnologie - Vrije Universiteit Brussel, B-1050 Brussels, Belgium; Brussels Center for Redox Biology, Vrije Universiteit Brussel, B-1050 Brussels, Belgium; Structural Biology Brussels, Vrije Universiteit Brussel, B-1050 Brussels, Belgium.
Cell Metab. 2020 Mar 3;31(3):642-653.e6. doi: 10.1016/j.cmet.2020.02.003.
Hydrogen peroxide (HO) is a key redox intermediate generated within cells. Existing probes for HO have not solved the problem of detection of the ultra-low concentrations of the oxidant: these reporters are not sensitive enough, or pH-dependent, or insufficiently bright, or not functional in mammalian cells, or have poor dynamic range. Here we present HyPer7, the first bright, pH-stable, ultrafast, and ultrasensitive ratiometric HO probe. HyPer7 is fully functional in mammalian cells and in other higher eukaryotes. The probe consists of a circularly permuted GFP integrated into the ultrasensitive OxyR domain from Neisseria meningitidis. Using HyPer7, we were able to uncover the details of HO diffusion from the mitochondrial matrix, to find a functional output of HO gradients in polarized cells, and to prove the existence of HO gradients in wounded tissue in vivo. Overall, HyPer7 is a probe of choice for real-time HO imaging in various biological contexts.
过氧化氢（HO）是细胞内产生的关键氧化还原中间体。现有的HO探针尚未解决超低浓度氧化剂检测的问题：这些报告分子不够灵敏、依赖pH值、亮度不足、在哺乳动物细胞中无功能或动态范围较差。在此，我们展示了HyPer7，首个明亮、pH稳定、超快且超灵敏的比率型HO探针。HyPer7在哺乳动物细胞和其他高等真核生物中具有完整功能。该探针由整合到脑膜炎奈瑟菌超灵敏OxyR结构域中的环状排列绿色荧光蛋白组成。利用HyPer7，我们得以揭示HO从线粒体基质扩散的细节，发现极化细胞中HO梯度的功能输出，并证明体内受伤组织中HO梯度的存在。总体而言，HyPer7是在各种生物学环境中进行实时HO成像的首选探针。
