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Conjugated dienoic acid peroxides as substrates in Chaetopterus bioluminescence system

Marine polychaete worm Chaetopterus variopedatus bioluminescence mechanism investigations date back several decades. Researchers from the Laboratory of Chemistry of Metabolic Pathways, the Laboratory of Biomolecular NMR-Spectroscopy and other subdivisions of the Institute of Bioorganic Chemistry RAS, in collaboration with scientists from the Laboratory of Photobiology of the IBP SB RAS, Moscow Institute of Physics and Technology, Pacific State Medical University, and Yeshiva University (USA) have established the structures of three substrates of the Chaetopterus luciferase bioluminescence reaction. The work was published in the International Journal of Molecular Sciences.

The glow of the marine polychaete worms Chaetopterus variopedatus was first described at the end of the 19th century, but its reaction mechanism remains unknown. The process referred to as bioluminescence, or the emission of light by living organisms, is common among the inhabitants of the sea depths. The Chaetopterus worm emits light through glowing mucus and short "flashes" of the posterior body segments. The first attempt to study the Chaetopterus bioluminescence was made by Osamu Shimomura (Nobel Prize in Chemistry 2008) and Frank Johnson, who used the 12th segment of the worm's body, containing glands that secrete luminous mucus. They demonstrated that five components are involved in Chaetopterus bioluminescence reaction: a dimeric protein with a molecular weight of 128–130 kDa (which they referred to as photoproteins), two cofactors (presumably a nucleoprotein and a lipid), hydrogen peroxide, and ferrous ions. Recent studies by our team have shown that the worm's bioluminescence reaction involves an enzyme (luciferase), a low molecular weight substrate (luciferin), and a cofactor - Fe2+ ions.

In order to determine the structure of the luciferin it was necessary to isolate it from a natural source, which proved impossible due to its low stability and low concentration in biomass. However, we resorted to another strategy, similar to the one tested earlier when studying the mechanism of fungal luminescence - to look for a bioluminescent substrate in other marine organisms. Substances exhibiting bioluminescent activity with C. variopedatus luciferase have been found in a number of organisms of both plant and animal origin (soft corals, cephalopods, algae). The greatest number of them, as it turned out, is contained in the algae Chaetomorpha linum. Ethanol extraction of the biomass and chromatographic separation of the supernatant made it possible to isolate three different components, respectively termed Hitik1, Hitik2, and Hitik3, displaying luminescent activity with Chaetopterus luciferase. The structures of the compounds were determined by NMR and MS analysis: it was shown that these are derivatives of unsaturated fatty acid peroxides. The structure of Hitik3 component was further confirmed by chemical synthesis. All obtained compounds, as well as other fatty acid and phospholipid peroxides, demonstrate bioluminescent activity with Chaetopterus luciferase extract in the presence of Fe2+ ions. Further study of the mechanism of bioluminescence may allow to develop applications for this process in the studies of one of the mechanisms of programmed cell death - ferroptosis.

Figure 1. Structures of bioluminescent active substrates Hitik1, Hitik2, Hitik3 with selective atom numbering. Line chart: Bioluminescence spectrum of synthetic Hitik3. Bar chart: Luminescent activity of chromatographic fractions of substrates (RLU) (lower right).

may 30, 2023