(801.12) Search and investigation of potential peptide agents of interaction between human organism and its microbiome

Poster Board Number: A214

Georgij Arapidi (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences), Anatoly Urban (Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency), Victoria Shender (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences), Alexandr Kuznetsov (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency), Olga Ivanova (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency), Leonid Lopuhov (Kazan Federal University), Alexander Laikov (Kazan Federal University), Nina Sharova (National Research Center – Institute of Immunology Federal Medical Biological Agency), Margarita Nikonova (National Research Center – Institute of Immunology Federal Medical Biological Agency), Alexandr Mitin (National Research Center – Institute of Immunology Federal Medical Biological Agency), Alexandr Martinov (National Research Center – Institute of Immunology Federal Medical Biological Agency), Tatiana Grigorieva (Kazan Federal University), Elena Ilina (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency), Vadim Ivanov (Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences), Vadim Govorun (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency)

Symbiotic interaction between the human body and its microbiota is an important issue of modern biomedicine and personalized medicine. However, little is known on molecular mechanisms of that relationship. Bearing in mind the ubiquitous participation of peptides in biomolecular interactions and regulatory processes we attempted direct search of blood peptides originated from microbial proteins. LC-MS/MS analysis was carried out of blood serum and plasma samples taken from 20 healthy donors on Q Exactive HF-X Hybrid Quadrupole-Orbitrap mass-spectrometer. Sample preparation was carried out based on our previously developed method of peptide desorption from the surface of major blood plasma proteins followed by standard chromatographic steps. Human microbiota protein sequences were taken from NIH Human Microbiome Project.

As a result, out of 13,625 identified peptides 912 were unique fragments of microbial precursors, which is about 6.69% of the total amount of detected bloodstream peptides. In 30 cases peptide identification was confirmed by mass-spectral study of individual synthetic samples. Absolute quantification by the mass-spectrometric method of multiple reaction monitoring (MRM) confirmed the presence of bacterial peptides in plasma and serum in the range of approximately 0.1 nM to 1 uM, which is comparable to physiologically significant hormone concentrations in human blood in normal conditions. The abundance of microbiota peptides reaches its maximum 5h after a meal. Most of the peptides correlate with the bacterial composition of the small intestine and are likely obtained by hydrolysis of membrane proteins with trypsin, chymotrypsin and pepsin — the main proteases of the gastrointestinal tract.

The isolated fraction of peripheral blood mononuclear cells showed increase secretion of proinflammatory cytokines, colony stimulating factors and chemoattractants as the response to the addition of some of the identified microbiotic peptides. Such peptides — identified both in the plasma and in the serum — have properties of resistance to fibrinolysis, since the pool of peptides is preserved after passing through the digestive tract. The physicochemical properties of the identified bacterial peptides are consistent with those required for the selective permeability of mucosal barriers. Our approach to the identification of microbiota peptides in the blood serum and plasma may be useful for determining the microbiota composition of hard-to-reach intestinal areas, such as the small intestine, and for monitoring the permeability of the intestinal mucosal barrier.

This work was supported by the Russian Science Foundation project no. 20-15-00400.