Ultra-sensitive sensor

The unique development of Russian scientists will allow researchers around the world to expand their understanding of the role of hydrogen peroxide in normal and pathological processes in living organisms

Artem Grigorov, IBH RAS Press Service

Hydrogen peroxide (H ₂ O ₂) is one of the most important biologically active molecules. It is a secondary product of aerobic metabolism, and is also generated by various enzyme systems. An abnormally high level of H ₂ O ₂ leads to the so-called oxidative stress, which is the cause of many pathological processes. In physiological concentrations, this compound performs intracellular and extracellular signaling functions, performs redox regulation of key biochemical processes. The regulatory potential of hydrogen peroxide is realized in extremely low nanomolar concentrations, the molecule has a high reactivity and, consequently, a short lifetime (up to several seconds). All together, this makes it very difficult to study the exact mechanisms of production and distribution of H ₂ O ₂ in cells and tissues, the physiological role of this molecule, its participation in pathological processes.

The development of domestic scientists has become the fruit of their many years of activity on the creation of biosensors of hydrogen peroxides. The first generation of such sensors, which at one time also received worldwide recognition and contributed to the expansion of knowledge in the field of redox biology, had, however, a number of disadvantages. The probes available up to now did not allow detecting ultra-low concentrations of H ₂ O ₂, that is, exactly those in which most of the functions of this molecule are realized. In addition, the sensors were dependent on the acidity (pH) of the medium and had insufficient brightness. Now researchers under the guidance of Doctor of Biological Sciences, Professor of the Russian Academy of Sciences Vsevolod Vadimovich Belousov, head of The Department of Metabolism and Redox Biology of the IBH RAS, managed to create the world's first hypersensitive reversible sensor H ₂ O ₂, stably operating in eukaryotic cells in a wide pH range. The work was published in the highly rated journal Cell Metabolism (Pak et al., Ultrasensitive Genetically Encoded Indicator for Hydrogen Peroxide Identifies Roles for the Oxidant in Cell Migration and Mitochondrial Function).

The HyPer7 biosensor consists of a fluorescent GFP protein crosslinked with the OxyR protein. Source: Vsevolod Belousov.

The new HyPer7 probe is a complex of green fluorescent protein integrated into the bacterial OxyR protein Neisseria meningitidis, sensitive to oxidation by hydrogen peroxide, but not by other active forms of oxygen. In order to develop it, scientists studied a variety of OxyR proteins in various bacteria, varied the scheme of integration of OxyR and fluorescent protein, and used directed and random mutagenesis. As a result, it was possible to obtain a unique visualization tool operating at ultra-low concentrations of hydrogen peroxide, with high brightness and short response time, independent of physiological pH changes.

The gradient of hydrogen peroxide in the cell sets the direction of its movement. The arrow shows the direction of movement. Source: Vsevolod Belousov, adapted from the original publication.

With the help of the developed sensor, the authors of the work have already managed to study the details of H ₂ O ₂ diffusion between mitochondria and cytosol, to investigate the role of hydrogen peroxide gradients in cell polarization, and in tissue damage. The development of Russian scientists will allow researchers around the world to visualize H ₂ O ₂ in various biological contexts and to investigate the role of this molecule in normal biochemical processes, as well as in a wide range of pathologies in which oxidative stress plays a key role: ischemic, neurodegenerative, inflammatory and oncological diseases.

Portal "Eternal youth" http://vechnayamolodost.ru