Basic research
Bacterial nanotube formation as a post mortem manifestation of a cell stress.

Authors: Barák, I., Muchová, K.

Bacterial nanotubes were discovered ten years ago. The unique abilities of these nanotubes have been attributed to processes such as the transfer of DNA, RNA and proteins between the cells of various bacteria, as well as the vampire-like sucking of nutrients from the eukaryotic cell. Our results are in contrast to previously published findings. We have shown that nanotubes in principle arise from every cell when we apply various stress factors, for example pressure or expose them to the action of antibiotics. The bacterial cell wall can maintain a pressure of up to twenty atmospheres inside the cell. However, if the wall is damaged either mechanically or by the activity of antibiotics, the cell can not withstand such a high pressure. This pressure will literally "blast" the cytoplasmic membrane in the form of a nanotube into the external environment through the holes created in the cell wall. An important finding was that the moment the cell "blasts" the nanotube, the cell is dying. This means that the formation of nanotubes is not a controlled biological process but a "post mortem" manifestation of a stressed cell.


Projects:

1. VEGA 2/0001/09 - Ako bunka nájde miesto asymetrického delenia počas sporulácie Bacillus subtilis. (How a cell finds a site of asymmetric division during sporulation of Bacillus subtilis.)
2. APVV-18-0104 – Asymetrické bunkové delenie počas tvorby bakteriálnej endospóry. (Asymmetric cell division during bacterial endospore formation.)

Publications:

1. POSPÍŠIL, J. - VÍTOVSKÁ, D. - KOFROŇOVÁ, O.- MUCHOVÁ, K.- ŠANDEROVÁ, H. - HUBÁLEK, M. - ŠIKOVÁ, M. - MODRÁK, M. - BENADA, O. - BARÁK, I. - KRÁSNY, L. Bacterial nanotubes as a manifestation of cell death. In Nature Communications, 2020, vol. 11, no. 4963. (2019: 12.121 - IF, Q1 - JCR, 5.569 - SJR, Q1 - SJR). 
2. MUCHOVÁ, K. - CHROMÍKOVÁ, Z. - BARÁK, I. Linking the peptidoglycan synthesis protein complex with asymmetric cell division during bacillus subtilis sporulation. In International Journal of Molecular Sciences, 2020, vol. 21, no. 4513. (2019: 4.556 - IF, Q1 - JCR, 1.317 - SJR, Q1 - SJR).
3. WOLLMAN, A. - MUCHOVÁ, K. - CHROMÍKOVÁ, Z. - WILKINSON, A.J. - BARÁK, I. - LEAKE, M.C. Single-molecule optical microscopy of protein dynamics and computational analysis of images to determine cell structure development in differentiating Bacillus subtilis. In Computational and Structural Biotechnology Journal, 2020, vol. 18, 1474–1486. (2019: 6.018 - IF, Q1 - JCR, 1.782 - SJR, Q1 - SJR).
4. GACEK-MATTHEWS, A. - CHROMÍKOVÁ, Z. - SULYOK, M. - LÜCKING, G. - BARÁK, I. - EHLING-SCHULZ, M. Beyond toxin transport: novel role of ABC transporter for enzymatic machinery of cereulide NRPS assembly line. In mBio, 2020, vol. 11,no. e01577. (2019: 6.784 - IF, Q1 - JCR, 3.876 - SJR, Q1 - SJR)
5. BODÍK, M. - KRAJČÍKOVÁ, D. - HAGARA, J. - MAJKOVA, E. - BARÁK, I. - ŠIFFALOVIČ, P. Diffraction pattern of Bacillus subtilis CotY spore coat protein 2D crystals. In Colloids and Surfaces B-Biointerfaces, 2020, vol. 197, 111425 (2019: 4.39 - IF, Q1 - JCR, Q1 - SJR)
   

Basic research
Pleiotropic key anti-sigma factor BldG is phosphorylated by several anti-sigma factors and regulates five sigma factors SigF, SigG, SigH, SigK, SigM in Streptomyces coelicolor A3 (2)

Authors: Ševčíková, B., Řežuchová, B., Mingyar, E., Homerová, D., Fecková, Ľ., Kormanec, J.

Streptomyces coelicolor contains up to 9 homologues of sigma factor SigB, 45 homologues of anti-sigma factor RsbW, 17 homologues of anti-anti-sigma factor RsbV, which is related to its differentiation. The anti-anti-sigma factor BldG plays an important role in differentiation and antibiotic production. It activates the sigma factors SigH and SigF through interaction with the anti-sigma factors UshX and RsfA, of which only RsfA phosphorylates BldG. BldG is phosphorylated in the rsfA mutant, indicating that other protein kinases phosphorylate BldG. We identified 15 RsbW homologues interacting with BldG, 7 of which phosphorylate BldG on Ser57. We investigated the interaction of these RsbW homologues with nine SigB factors. Only SCO7328 interacted with sigma factors SigG, SigK, SigM. These results suggest a unique phosphorylation mechanism of BldG regulation in the activation of five sigma factors, via multiple anti-sigma factors, suggesting a complex regulation of stress response and differentiation in S. coelicolor (figure below).


Projects:
VEGA 2/0026/20 – Signálne kaskády regulácie sigma faktorov RNA polymerázy pri odozve na stres, bunkovej a fyziologickej diferenciácii u pôdnych baktérií rodu Streptomyces (Signaling cascades of RNA polymerase sigma factor regulation in response to stress, cell and physiological differentiation in soil bacteria of the genus Streptomyces)
Publications:
ŠEVČÍKOVÁ, B. - REŽUCHOVÁ, B. - MINGYAR, E. - HOMEROVÁ, D. - NOVÁKOVÁ, R. - FECKOVÁ, Ľ. - KORMANEC, J. Pleiotropic anti-anti-sigma factor BldG is phosphorylated by several anti-sigma factor kinases in the process of activating multiple sigma factors in Streptomyces coelicolor A3(2). In Gene, 2020, vol. 755, no. 144883. (2019: 2.984 - IF, Q2 - JCR, 0.898 - SJR, Q1 - SJR)
 

Basic research
Identification of a new subfamily in the α-amylase family GH13 represented by α-amylase from the halophilic archaeon Haloarcula hispanica

Authors: Janeček, Š., Zámocká, B.

α-Amylase catalyzes the endohydrolysis of α-1,4-glucosidic bonds in starch and related α-glucans. In the CAZy database (http://www.cazy.org/), most α-amylases are classified into the GH13 glycoside hydrolase family, which currently contains more than 107 000 sequences and more than 30 different enzyme specificities. Most members of the family have so far been divided into 42 subfamilies; other subfamilies are established continuously. Based on a detailed bioinformatic study, a new subfamily represented by an α-amylase from the halophilic archaeon Haloarcula hispanica was proposed, covering a group of hypothetical haloarchaeal α-amylases. The group shares unique sequence similarities with each other and forms its own cluster in the evolutionary tree, separated from representatives of already established GH13 subfamilies. From an evolutionary point of view, this new subfamily is most closely related to the two GH13 clusters of α-amylase-specific subfamilies, i.e. to subfamilies GH13_5, 6 and 7, as well as GH13_15, 24, 27 and 28.


Projects:
VEGA 2/0146/17 – Evolúcia amylolytických enzýmov (Evolution of amylolytic enzymes)
Publications:
JANEČEK, Š. - ZÁMOCKÁ, B. A new GH13 subfamily represented by the α-amylase from the halophilic archaeon Haloarcula hispanica. In Extremophiles, 2020, vol. 24, 207–217. (2019: 2.462 - IF, Q3 - JCR, 0.734 - SJR, Q2 - SJR)

Applied research
New antimicrobial recombinant protein EN534-C with lytic properties against pathogenic strains of Streptococcus agalactiae.

Authors: Bukovská, G., Bocánová, L., Halgašová, N., Kajsiková, M.

Pathogenic bacteria Streptococcus agalactiae group B (GBS) cause serious diseases in humans and are known mainly as the cause of infections in pregnant women and in children. We have prepared a novel endolysin EN534-C, which is the first endolysin derived from the Streptococcus agalactiae KMB-534 (GBS) human clinical isolate with an arrangement of amidase-3, CHAP and LysM domains. We overproduced the recombinant endolysin in the E. coli expression system, optimized the protein isolation conditions and the reaction conditions for determining the lytic activity. We tested endolysin EN534-C on a spectrum of bacterial substrates from cell walls or living cells of streptococcal strains. We confirmed its specific action against strains of streptococci, especially against the pathogenic bacterium S. agalactiae GBS. The new endolysin EN534-C has potential use for the treatment, prevention and diagnosis of diseases caused by pathogenic streptococcal bacteria and represents an alternative to antibiotics, especially in cases of pregnant women and newborns.
Projects:
APVV-16-0168 - Príprava bakteriofágov na terapiu vaginálnych a močových infekcií (Preparation of bacteriophages for the treatment of vaginal and urinary tract infections)
Publications:

LICHVARIKOVÁ, A. - ŠOLTYS, Katarína - SZEMEŠ, T. - SLOBODNÍKOVÁ, L. - BUKOVSKÁ, G. - TURŇA, J. - DRAHOVSKÁ, H. Characterization of Clinical and Carrier Streptococcus agalactiae and Prophage Contribution to the Strain Variability. In Viruses, 2020, vol. 12, p. 1323. (2019: 3.816 - IF, Q2 - JCR, 1.633 - SJR, Q1 - SJR).

Nová patentová prihláška č. PP 50075-2020 v Slovenskej republike, Prihlasovatelia: Ústav molekulárnej biológie SAV, Univerzita Komenského v Bratislave, Názov: "Antimikrobiálny proteín, antimikrobiálny rekombinantný proteín s lytickými vlastnosťami, expresný vektor, spôsob ich prípravy a použitie“.

(New patent application no. PP 50075-2020 in the Slovak Republic, Applicants: Institute of Molecular Biology of the Slovak Academy of Sciences, Comenius University in Bratislava, Title: "Antimicrobial protein, antimicrobial recombinant protein with lytic properties, expression vector, method of their preparation and use".)

 

Applied research
Application of third generation sequencing (MinION, Oxford Nanopore Technologies) to study the microbiota of cultural heritage samples.

Authors: Pangallo, D., Bučková, M., Puškárová, A., Kraková, L., Planý, M.

Our group is probably the first [1] or among the first groups in the world to use MinION sequencing to investigate the microbiome of several samples of a historical carnauba wax seal (XVIII century) and funerary accessories [2] (XVII century). High-throughput sequencing analysis showed the presence of a complex representation of microorganisms on the carnauba wax seal, consisting mainly of representatives of microscopic filamentous fungi that showed interesting properties related to lipid and lead processing. The formation of lead soaps and secondary biogenic minerals was also described. In addition, this technique has been useful, in the context of the investigation of funerary accessories, for the detection of dangerous degrading microorganisms that are able to hydrolyze various organic substrates such as fibroin, keratin and cellulose. Bacteria responsible for metal corrosion and biomineralization and entomopathogenic and phytopathogenic fungi were also discovered.


Projects:

1. VEGA 2/0061/17 – Inovatívne stratégie dezinfekcie: vplyv esenciálnych olejov na mikroflóru a materiály objektov kultúrneho dedičstva (Innovative disinfection strategies: the impact of essential oils on microflora and heritage materials)
2. APVV-19-0059 – Farebné škvrny na historických papieroch: biologická a chemická charakterizácia spojená s ich odstraňovaním (Colored stains on historical papers: biological and chemical characterization associated with their removal)

Publications:

1. ŠOLTÝS, K. - PLANÝ, M. - BIOCCA, P. - VIANELLO, V. - BUČKOVÁ, M. - PUŠKÁROVÁ, A. - SCLOCCHI, M.C. - COLAIZZI, P. - BICCHIERI, M. - PANGALLO, D.** - PINZARI, F. Lead soaps formation and biodiversity in a XVIII. In Environmental microbiology, 2020, vol. 22, 1517–1534. (2019: 4.933 - IF, Q1 - JCR, 2.180 - SJR, Q1 - SJR).
2. KISOVÁ, Z. - PLANÝ, M. - PAVLOVIĆ, J. - BUČKOVÁ, M. - PUŠKÁROVÁ, A. - KRAKOVÁ, L. - KAPUSTOVÁ, M. - PANGALLO, D. - ŠOLTYS, K. Biodeteriogens Characterization and Molecular Analyses of Diverse Funeral Accessories from XVII Century. In Applied Sciences-Basel, 2020, vol. 10, p. 5451. (2019: 2.474 - IF, Q2 - JCR, 0.418 - SJR, Q1 - SJR).
   

International research projects
MAL protein (myelin and lymphocyte protein) as a functional indicator of CD4 T-cell differentiation and activation

Authors: Leksa, V.

In an international collaboration, we identified the MT3 antigen as the MAL protein, which is known as a transport receptor for the Lck kinase. It is showing high surface expression on naïve CD4+ T-cells and reduced expression on effector memory T-cells. Our detailed experiments showed that regulation of MAL protein expression in T-cells is associated with activation-induced differentiation of human T-cells but not with membrane localization of Lck signaling capacity.


Projects:

1. Austrian Science Fund (P19014-B13 a P22908)
2. APVV-16-0452 - Regulácia pericelulárnej proteolýzy: od molekulárnych mechanizmov k novým subsetom imunitných buniek a terapeutickým nástrojom (Regulation of pericellular proteolysis: from molecular mechanisms to new immune cell subsets and therapeutic tools)
3. VEGA 2/0020/17 - Ľudský mliečny bioaktívny glykoproteín laktoferín ako regulátor homeostázy (Human milk bioactive glycoprotein lactoferrin as a regulator of homeostasis)

Publications:
LEITNER, J. - MAHASONGKRAM, K. - SCHATZLMAIER, P. - PFISTERER, K. - LEKSA, V.- PATA, S. - KASINRERK, W. - STOCKINGER, H.** - STEINBERGER, P.**. Differentiation and activation of human CD4 T cells is associated with a gradual loss of myelin and lymphocyte protein. In European Journal of Immunology, 2020, vol. 51, p. 848-863. (2020: 5.532 - IF, Q2 - JCR, 2.272 - SJR, Q1 - SJR)