Lab of Proteome Research

About our Work

About our Work

The laboratory was founded in 2005. The aim of the laboratory was provision of services for other laboratories in the field of mass-spectrometry of proteins, peptides and other high-molecular compounds. A year later the laboratory initiated its own research of Mollicutes bacteria with reduced genome. Currently the scope of studied organisms include bacteria Mycoplasma gallisepticum, Spiroplasma meliferum and Acholeplasma laidlawii, Helicobacter pylori and Eshcerichia coli.

After 11 years of successful work the laboratory expanded the range of methods and technologies, which allowed it to carry out complex studies of principles of biological systems organization and function on the systemic level including genome, transcriptome, proteome and metabolome.

Currently a number of scientific projects have been accomplished:

  • Transcriptional and proteomic profiling of a set of clinical strains of Helicobacter pylori was carried out.
  • The model of intracellular infection of eukaryotic cells with Mycoplasma gallisepticum was developed. Proteome profiling of gallisepticum during intracellular infection was carried out. It was demonstrated that M. gallisepticum proteome switches to a new stable attractor after infection, which can self-sustain for long time.
  • The map of epigenetic modifications of Mycoplasma gallisepticum was obtained using PacBio sequencing.
  • Proteome and ribosome profiling of Mycoplasma gallisepticum in stress was carried out. The principles of stress response in mycoplasma were identified.
  • Transcriptional profiling of Mycoplasma gallisepticum, Spiroplasma meliferum and Acholeplasma laidlawii was carried out. Respective promoter maps were obtained and the structures of promoters in respective species were identified. Transcription control circuits were identified. The absolute copy number of mRNAs per cell was identified for gallisepticum.
  • Metabolome analysis of Mycoplasma gallisepticum, Spiroplasma meliferum and Acholeplasma laidlawii was carried out.
  • Metabolic reconstruction of Mycoplasma gallisepticum, Spiroplasma meliferum and Acholeplasma laidlawii was carried out.
  • A set of tools for genetic engineering was developed for Mycoplasma gallisepticum.
  • A proteome content of minimal cell was inferred using comparative proteome analysis of Mollicutes.
  • The model of dormant state of Mycoplasma gallisepticum was developed. The model includes cultivation on depleted medium under low temperature. Proteome profiling of the obtained nanoforms and recultivated forms was carried out.
  • The genome of Mycoplasma gallisepticum S6 strain was sequenced.


There are two groups in the laboratory who study systems biology of mycoplasmas and metaproteomics of gut disorders including Crohn's disease.

Group of Systems Biology of Mycoplasmas
Group of Metaproteome Analysis

Meet our Team

Meet our Team

Lab of Proteome Research

Gleb Fisunov, PhD, Head of laboratory

Daria Rakitina, PhD, Senior researcher

Olga Pobeguts, PhD, Senior researcher

Anna Vanyushkina, PhD, Senior researcher

Valentina Ladygina, PhD, Researcher

Tatiana Semashko, PhD, Researcher

Tatiana Gribova, PhD, Researcher

Yulia Baikova, PhD, Researcher

Daria Matyushkina, Junior Researcher

Mark Levites, Junior Researcher

Natalia Zakharzhevskaya, Junior Researcher

Ivan Butenko, Junior Researcher

Maria Galyamina, Junior Researcher

Valeria Orlova, PhD, Technician

Irina Garanina, PhD student

Daria Evsyutina, PhD student

Olga Bukato, PhD student

Group of Systems Biology of Mycoplasmas

Group of Systems Biology of Mycoplasmas


Mycoplasmas are bacteria of class Mollicutes. They are specialized branch of microorganisms related to Gram-positive bacteria. The most of Mollicutes are obligate parasites. However, there are facultative parasites and commensals. Mollicutes feature a number of unusual properties including significant genome reduction and lack of cell wall. Many species feature motility based on mechanisms unique among bacteria.

Mycoplasma gallisepticum is an avian respiratory pathogen. It is a close relative of human pathogens M. pneumonia, M. genitalium and Ureaplasma spp. M. gallisepticum is a typical mycoplasma. Its genome size is near 1 Mb. M. gallisepticum (like many species) features a specialized attachment (or terminal) organelle and motility apparatus associated with it. About 10% of M. gallisepticum genome codes surface hemagglutinins. Respective genes are organized into cassettes. The repertoire of expressed hemagglutinins may vary during the development of infection, which provides the basis of immune escape. The respective underlying mechanisms are not yet fully understood.

Spiroplasma melliferum has two hosts: flower plants and honeybee. The lifecycle of spiroplasma includes successive change of hosts which involves the change of physiology and morphology of the spiroplasma cells. This species is among few Mollicutes with extrachromosomal elements.

Acholeplasma laidlawii is a free-living organism. Its ecological niche is not clear and most likely it is a saprophyte of aquatic sediment. It probably may be a facultative pathogen of plants. This bacterium takes intermediate position between “normal” bacteria and reduced mycoplasmas. It has complex metabolism and broad regulatory networks, however still featuring key features of mycoplasmas like reduced genome, low GC-content, absence of cell wall and terminal organelle.

Mycoplasmas are members of the class Mollicutes. Mollicutes represent a group of highly specialized microorganisms relating to the Gram-positive bacteria. Most of the Mollicutes are obligate parasites, although facultative parasites and commensals are also present. Mollicutes have distinctive features such as considerably reduced genome and lack of a cell wall. Many of the mycoplasmas are unique for their mechanism of locomotion which is not found in other bacteria.

Mycoplasma gallisepticum is a pathogen causing a chronic respiratory disease in birds. M. gallisepticum is a relative of human pathogens such as M. pneumonia, M. genitalium, and members of the genus Ureaplasma. It may therefore serve as a model system for studies on these human infections. M. gallisepticum is a “classic” mycoplasma. The genome size for M. gallisepticum is about 1 million base pairs. M. gallisepticum possesses a specialized terminal organelle that enables the bacterium to attach to host cells as well as provides a gliding mechanism which may be described by the so-called centipede model. An interesting feature of M. gallisepticum is that the cassettes of surface antigens account for about 10% of its genome. These proteins play two roles: they provide adherence to host cells and contribute to the bacterial evasion of the immune response. The latter proceeds via the replacement of the cassette’s antigen that elicited the immune response to another antigen. Mechanism of this replacement is not clear yet.

Spiroplasma melliferum which parasitizes on bees and flowering plants is distinguished by its life cycle that includes migration from one type of the host to another. Change of a host is associated with physiological and morphological changes. S. melliferum is one of the few mycoplasmas that carry extrachromosomal elements.

Acholeplasma laidlawii  is a free living microorganism. This bacterium is of particular interest because it occupies an intermediate position between normal bacteria and classic mycoplasmas due to a number of physiological features.

Current projects

The main problem of our research on systems biology of mycoplasmas is mutual interaction between regulation and chaos in the cell that ultimately give rise to a stable system. It was discovered that mycoplasmas are prone to noise and most of its response to a given condition may represent noise that has nothing to do with adaptation. In other conditions mycoplasma may demonstrate alternative stable state (it may be termed attractor or cell’s memory) on the level of proteome with little or no change on the level of transcription. Proteome self-stability may represent a novel layer of regulation beyond transcription-factor based systems.

The objective of our work is to investigate a stress adaptive mechanism in Mycoplasma gallisepticum using models of oxidative, osmotic, and heat stress. Because of the parasitic lifestyle of Mycoplasma gallisepticum, these stresses are physiologically relevant kinds of stressful conditions to which bacteria are subject due to host’s immune response. Our project aimed to conduct a full systematic investigation of all levels of adaptation of this bacterium. Our approach included genome sequencing, creation of stress models, transcriptomic analysis in different stress states, and proteomic analysis of the chosen states. The transcriptome profiling also involved mapping of functional elements, including promoters, operons, and non-coding RNAs. Mapping of the transcription initiation sites allowed us to create a model of a M. gallisepticum promoter. New genes involved in response to stress were found. It was shown that heat stress leads to the loss of M. gallisepticum surface antigens, which may be an element of bacterial defense against host’s immune response.

Interaction of mycoplasmas with eukaryotic cells

The long-term search for gene expression control in mycoplasmas resulted in the general understanding that it is mostly absent. However, a widely used approach which implies testing of an organism under a set of stress laboratory conditions does not seem to be the best choice for mycoplasmas. These bacteria evolved towards tight interaction with the host and respective mechanisms may not be revealed under artificial conditions in many cases. Contrary to what was seen under standard stress conditions (heat, hyperosmotic and oxidative stresses) mycoplasma demonstrates complex reorganization of the proteome during intracellular infection. Proteomic changes affected a broad range of processes including metabolism, glycolysis, translation and oxidative stress response. The obtained data led to conclusion that there are different stable states of the proteome (attractors) that are capable of self-sustain during significant time when cells are returned to the cultural medium. The mechanisms of this phenomenon are not yet understood. The respective research may shed light on the basic properties of self-organization of living systems.

Identification of regulatory mechanisms in mycoplasmas

Mycoplasmas can be treated as a close approximation to “minimal cell” – a cell with a minimal set of genes capable of self-replication on a medium. General reduction of mycoplasmas involves reduction of gene expression control systems as well. Only few transcription factors and other regulators are known for them and even less have known function. The study of gene expression regulation in mycoplasmas aims to solve two problems. First, which processes require regulation of the level of gene expression in the minimal cell? Second implies prediction of cell’s response to the given condition based on the known regulatory network.

Group of Metaproteome Analysis

Group of Metaproteome Analysis

Current projects

Analysis of major proteins in microbiota of various parts of healthy or inflamed human intestine

The aim of this project is to detect specific alterations in major proteins of microbiota of human intestine during inflammation. It will suggest possible associations between certain disease and alterations in composition and functionality of microbiota.

Basic method used in this project: full metaproteome analysis of proteins extracted from feces, biopsy, or ileal content (Ultimate 3000 HPLC(Dionex, Thermo); nano-LC-QTOF-MS-MS, maxis, Bruker).

Intestinal microbiota of patients with Crohn’s disease

The aim of this project is to study certain stages of Crohn's disease (CD) pathogenesis. CD was observed to develop as a complication of massive medical interventions. Such interventions are liver and renal transplantations (accompanied by immunosuppressive therapy) and long antibiotics treatment (as a result of heavy infections or trauma). Therefore, understanding of CD-inducing mechanism can drive to improvement of treatment and rehabilitation protocols, preventing complications and reducing duration of rehabilitation period.

Crohn's disease is a chronic intestine inflammation. Generally, chronic character of the disease severely reduces the quality of patient’s live, often driving to invalidization. Its pathogenesis mechanism is yet unclear. The most accepted mechanism is the provocation of unregulated immune response by opportunistic microbiota, colonising intestinal mucosa. Escherichia coli is regarded as the most probable pathogen or causative agent (CD-associated, adherent-invasive E. coli), since 30-60% of CD-patients demonstrate its increased content in intestine microbiota (here literature data is supported by our observation). Complex and throughout characteristic of CD-associated E. coli can provide more effective methods for rehabilitation of these patients. Revealing properties that define CD-associated E. coli (genes, proteins, metabolic pathways etc) might give new targets for therapy. Comparative analysis of antibiotics and phage resistance, adherent invasive abilities, biofilm formation and effect of inflammation agents (cell, protein and their products) will shed a light on interaction of E. coli and human host on different stages of infection.

Basic methods used in this project: biotyping of bacteria (Biotyper, MALDI-mass spectrometer microflex, Bruker), 2D-electroforesis peptide mass fingerprinting (MALDI/TOF autoflex, Bruker), full proteome analysis (Ultimate 3000 HPLC(Dionex, Thermo); nano-LC-QTOF-MS-MS, maxis, Bruker).



  • Laminar-flow cabinets, thermostats, CO2-incubators for cultivation of bacterial and eukaryotic cells.
  • Equipment for 1D and 2D electrophoresis, isoelectrofocusing and Western-blotting.
  • Real-time PCR machine BioRad CFX96.
  • Typhoon imager.
  • MS and LC-MS machines: AB SCIEX TripleTOF® 5600 System, AB SCIEX QTRAP® 4500 LC/MS/MS System, Bruker Daltonics MaXis, Agilent 1100 LC/MSD Trap SL, Ultraflex Tof-Tof (MALDI)




  1. Fisunov G.Y., Garanina I.A., Evsyutina D.V., Semashko T.A., Nikitina A.S., Govorun V.M. Reconstruction of transcription control networks in Mollicutes by high-throughput identification of promoters. Frontiers in Microbiology; 2016; doi: 10.3389/fmicb.2016.01977
  2. Matyushkina D., Pobeguts O., Butenko I., Vanyushkina A., Anikanov N., Bukato O., Evsyutina D., Bogomazova A., Lagarkova M., Semashko T., Garanina I., Babenko V., Vakhitova M., Lagygina V., Fisunov G., Govorun V. Phase Transition of the Bacterium upon Invasion of a Host Cell as a Mechanism of Adaptation a Mycoplasma gallisepticum model. Scientific Reports; 2016; 1–13. doi: 10.1038/srep35959.
  3. Fisunov G.Y., Evsyutina D.V., Garanina I.A., Arzamasov A.A., Butenko I.O., Altukhov I.A., Nikitina A.S., Govotun V.M. Ribosome profiling reveals an adaptation strategy of reduced bacterium to acute stress. Biochimie; 2017; 132:66–74. doi: 10.1016/j.biochi.2016.10.015
  4. Fisunov G.Y., Evsyutina D.V., Semashko T.A., Arzamasov A.A., Manuvera V.A., Letarov A.V., Govorun V.M. Binding site of MraZ transcription factor in Mollicutes. Biochimie; 2016; 125:59–65. doi:10.1016/j.biochi.2016.02.016.
  5. Vanyushkina A, Fisunov GY, Gorbachev AY, Kamashev DE, Govorun VM, Metabolomics analysis of three Mollicute speciesPLoS One2014, 9(3):e89312, doi: 10.1371/journal.pone.0089312
  6. Mazin PV, Fisunov GY, Gorbachev AY, Kapitskaya KY, Altukhov IA, Semashko TA, Alexeev DG, Govorun VM, Transcriptome analysis reveals novel regulatory mechanisms in a genome-reduced bacterium, Nucleic Acids Research, 2014, 42(21):13254-68, doi: 10.1093/nar/gku976
  7. Gorbachev AY, Fisunov GY, Izraelson M, Evsyutina DV, Mazin PV, Alexeev DG, Pobeguts OV, Gorshkova TN, Kovalchuk SI, Kamashev DE, Govorun VM., DNA repair in Mycoplasma gallisepticum, BMC Genomics,  14:726. 2013. doi: 10.1186/1471-2164-14-726
  8. Tyakht AV, Kostryukova ES, Popenko AS, Belenikin MS, Pavlenko AS, Larin AK, Karpova IY, Selezneva OV, Semashko TA, Ospanova EA, Babenko VV, Maev IV, Cheremushkin SV, Kucheryavy YA, Shcherbakov PL, Grinevich VB, Efimov OI, Sas EI, Abdulkhakov RA, Abdulkhakov SA, Lyalyukova EA, Livzan EA, Vlassov VV, Sagdeev RZ, Tsukanov VV, Osipenko MF, Kozlova IV, Tkachev AV, Sergienko VI, Alexeev DG., Govorun V.M., Novel human gut microbiota compositions found across urban and rural populations of Russia. Nature communications,2013 Sep 16;4:2469. doi: 10.1038/ncomms3469.
  9. Zgoda VG, Kopylov AT, Tikhonova OV, Moisa AA, Pyndyk NV, Farafonova TE, Novikova SE, Lisitsa AV, Ponomarenko EA, Poverennaya EV, Radko SP, Khmeleva SA, Kurbatov LK, Filimonov AD, Bogolyubova NA, Ilgisonis EV, Chernobrovkin AL, Ivanov AS, Medvedev AE, Mezentsev YV, Moshkovskii SA, Naryzhny SN, Ilina EN, Kostrjukova ES, Alexeev DG, Tyakht AV, Govorun VM, Archakov AI. Chromosome 18 transcriptome profiling and targeted proteome mapping in depleted plasma, liver tissue and HepG2 cellsJ Proteome Res. 2013 Jan 4;12(1):123-34. doi: 10.1021/pr300821n.
  10. Lazarev VN, Shkarupeta MM, Polina NF, Kostrjukova ES, Vassilevski AA, Kozlov SA, Grishin EV, Govorun VM. Antimicrobial peptide from spider venom inhibits Chlamydia trachomatis infection at an early stageArch Microbiol2013 Mar;195(3):173-9. doi: 10.1007/s00203-012-0863-5.
  11. Naumov VA, Generozov EV, Zaharjevskaya NB, Matushkina DS, Larin AK, Chernyshov SV, Alekseev MV, Shelygin YA, Govorun VM. Genome-scale analysis of DNA methylation in colorectal cancer using Infinium HumanMethylation450 BeadChipsEpigenetics2013 Jul 17;8(9), 921–934.
    doi: 10.4161/epi.25577
  12. Kharlampieva DD, Manuvera VA, Podgornyi OV, Kovalchuk SI, Pobeguts OV, Altukhov IA, Alexeev DG, Lazarev VN., Govorun VM., Purification and characterisation of recombinant Bacteroides fragilis toxin-2. 
    Biochimie, 2013, 95(11): 2123–2131.
    Aug 15. doi:pii: S0300-9084(13)00273-3. 10.1016/j.biochi.2013.08.005.
  13. Osterman IA, Ustinov AV, Evdokimov DV, Korshun VA, Sergiev PV, Serebryakova MV, Demina IA, Galyamina MA, Dontsova OA., Govorun VM.
    A nascent proteome study combining click chemistry with 2DE. Proteomics, 13 (1), 17-21, 2013. doi 10.1002/pmic.201200393
  14. Alexeev D, Kostrjukova E, Aliper A, Popenko A, Bazaleev N, Tyakht A, Selezneva O, Akopian T, Prichodko E, Kondratov I, Chukin M, Demina I, Galyamina M, Kamashev D, Vanyushkina A, Ladygina V, Levitskii S, Lazarev V., Govorun V., Application of Spiroplasma melliferum proteogenomic profiling for the discovery of virulence factors and pathogenicity mechanisms in host-associated spiroplasmas. J Proteome Res., 11 (1), 224-36, 2012.  doi: 10.1021/pr2008626
  15. Sergiev PV, Lesnyak DV, Burakovsky DE, Svetlov M, Kolb VA, Serebryakova MV, Demina IA, Dontsova OA, Bogdanov AA., Govorun VM, Non-stressful death of 23S rRNA mutant G2061C defective in puromycin reaction.
    J Mol Biol., 416 (5), 656-67, 2012. doi:10.1016/j.jmb.2012.01.005
  16. GY. Fisunov, DG. Alexeev, NA. Bazaleev, VG. Ladygina, MA. Galyamina, IG. Kondratov, NA. Zhukova, MV. Serebryakova, IA. Demina, Govorun V.M., Core Proteome of the Minimal Cell: Comparative Proteomics of Three Mollicute Species, PLoS One. 6(7), e21964, 2011. doi: 10.1371/journal.pone.0021964
  17. Kamashev D, Oberto J, Serebryakova M, Gorbachev A, Zhukova Y, Levitskii S, Mazur AK, Govorun VM., Mycoplasma gallisepticum Produces a Histone-like Protein That Recognizes Base Mismatches in DNA. Biochemistry, 93 (7), 1102-9, 2011. doi: 10.1021/bi2009097
  18. Lazarev VN, Levitskii SA, Basovskii YI, Chukin MM, Akopian TA, Vereshchagin VV, Kostrjukova ES, Kovaleva GY, Kazanov MD, Malko DB, Vitreschak AG, Sernova NV, Gelfand MS, Demina IA, Serebryakova MV, Galyamina MA, Vtyurin NN, Rogov SI, Alexeev DG, Ladygina VG, Govorun VM. Complete genome and proteome of Acholeplasma laidlawiiJ Bacteriol., 193 (18), 4943-53, 2011. doi: 10.1128/JB.05059-11.
  19. Levitskiy SA, Sycheva AM, Kharlampieva DD, Oberto J, Kamashev DE, Serebryakova MV, Moshkovskii SA, Lazarev VN, Govorun V.M., Purification and functional analysis of recombinant Acholeplasma laidlawii histone-like HU protein. Biochimie, 93 (7), 1102-9, 2011. doi:10.1016/j.biochi.2011.03.005
  20. Serebryakova MV, Demina IA, Galyamina MA, Kondratov IG, Ladygina VG, Govorun V.M., The acylation state of surface lipoproteins of mollicute Acholeplasma laidlawiiJBC, 286(26), 22769-76, 2011.  doi: 10.1074/jbc.M111.231316.
  21. Демина И.А., Серебрякова М.В., Ладыгина В.Г., Галямина М.А., Жукова Н.А., Алексеев Д.Г., Фисунов Г.Ю., Говорун В.М., Сравнительная протеомная характеристика микоплазм (молликут)Биоорганическая химия, Т. 37, №1, 70–80, 2011.
  22. Momynaliev KT, Kashin SV, Chelysheva VV, Selezneva OV, IA Demina Serebryakova MV, Alexeev D, Ivanisenko VA, Aman E., Govorun VM., Functional divergence of Helicobacter pylori related to early gastric cancer, J Proteome Res. 9(1), 254-67, 2010. DOI: 10.1021/pr900586w
  23. Lazarev VN, Borisenko GG, Shkarupeta MM, Demina IA, Serebryakova MV, Galyamina MA, Levitskiy SA, Govorun VM. The role of intracellular glutathione in the progression of Chlamydia trachomatis infection. Free Radic Biol Med. 49(12), 1947-55, 2010. doi: 10.1016/j.freeradbiomed.
  24. Golovina AY, Sergiev PV, Golovin AV, Serebryakova MV, Demina I, Dontsova OA., un VM., The yfiC gene of E. coli encodes an adenine-N6 methyltransferase that specifically modifies A37 of tRNA1Val(cmo5UAC), RNA, 15(6), 1134-41, 2009. doi: 10.1261/rna.1