The glutamine signaling in bacteria: molecular mechanisms of nitrogen metabolism regulation and signaling in Firmicutes
RFBR-15-04- 02583a
Project supervisor: A. Kayumov
The aim of the project is the investigation and elucidation of molecular mechanisms of nitrogen
metabolism regulation of Firmicutes. Despite of wide application of these bacteria in various biotechnological processes like silage production, lactic-acid products, probiotics and Bacillus и Lactobacillus-based drugs against disbacteriosis, the nitrogen metabolism of these bacteria isnot fully studied. The most prokaryotes assimilate nitrogen in the reduced form. Some species use inorganic nitrogen (ammonium, nitrates, molecular nitrogen) or simple forms of organic nitrogen (urea). All forms of nitrogen are firstly converted to the ammonium, which can be easily transformed into an amino group. The main role belongs to enzyme glutamine synthetase which produces glutamine from glutamic acid and ammonia. Glutamine is a donor of the amino group for the biosynthesis of the other organic compounds of the cell. Therefore, the intracellular level of glutamine availability serves as a signal of nitrogen availability for bacteria. In bacilli and lactic acid bacteria the nitrogen metabolism is controlled by transcription factors TnrA and GlnR. Interesting, TnrA does not have any sites of covalent modification or interaction with effector molecules. Its activity is believed to be regulated by interactions with glutamine synthetase and GlnK protein. In lactobacilli the regulation of nitrogen metabolism
remains unexplored.
Development of complex inhibitors of bacterial biofilms on the temporary and chronic implants based on
the furanones derivatives and immobilized enzymes
RNF 15-14-00046
Project supervisor: A. Kayumov
The replacement of damaged or removed tissue by the artificial implants is 
commonly required in clinical practice. Due to the contact with the outer skin, the surrounding surface becomes the main "gateway" for bacteria, which increases the inflammation risk. Many human resident bacteria such as Micrococcus sp, Staphylococcus sp are largely presented on the surface of human and animal skin but normally do not cause inflammation. However, getting into the inner tissues, they activate the multiple pathological factors (such as factors invasion, adhesion, and aggression) and form biofilms on implants and catheters. Being in biofilm, bacteria become resistant to the immune system and cause chronic inflammation.
The polymicrobial biofilms as a patogenicity factor: their modelling and identification of molecular targets for drug design
The project is addressing the complex problem of prevention of the infectious complications 
during and after the installation of implants (vascular, bone, teeth, etc), electrodes and catheters. In particular, this problem is associated with the formation of bacterial biofilms by opportunistic and pathogenic microflora on non-immunogenic artificial surfaces, which leads to the development of chronic inflammation and, as a consequence, increases of regeneration time, assimilation and rejection of the implant element, and development of sepsis.
The statistical analysis of biomolecules
The project is expected to develop methods for rapid analysis of comparative metagenomes and synthetic (-translated) metaproteoms of bacterial communities to answer foundamental questions of biological diversity, health and environmental monitoring habitats using modern methods of statistical physics without the u reconstruction and assembly of genomic sequences.
Navadays, because of the development of next-generation sequencing techniques a large database of bacterial metagenomes from the different habitats are publicly available. Their analysis is mainly performed by classical multiple alignment, sometimes with subsequent assembly of genomes and their comparative analysis. However, this approach is very resource-intensive and at the same time allows to annotate a relatively small portion of the information contained in the empirical metagenome, particularly characterized by high biodiversity. First, the incomplete genomes of rare species in the sample does not allow to reconstruct their genomes. Secondly, a significant obstacle to the reconstruction portion genomes can also be the presence of variable regions of the genetic code. These circumstances cause the need for parallel development of alternative methods for the rapid analysis of metagenomic information that would solve a number of problems without full or partial reconstruction of the metagenome.
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Development of new biotechnological strains of Lactobacillus
The main focus of our research is to study the probiotic
properties of Lactobacillus, in particular, to identify the functional role of special physiological characteristics and properties of lactobacilli in their probiotic activity. Lactobacillus plantarum was demonstrated to produce regulatory molecule nitric oxide (NO) via NO-synthase (NOS) pathway similar to mammalian cells [Yarullina et al 2015// JMRR]. We also showed the activation of bacterial NO-synthase by NOS substrate L-arginine and an increased level of NO production in lactobacilli under stress conditions, for example, high temperature [Smolentseva et al 2012// Mol Gen Microbiol Virol (Rus)]. The biological role of microbial NO in bacteria and macroorganisms is under research. We have shown that lactobacilli may influence through nitric oxide on protective and adaptive plant responses to dehydration caused by oxidative stress [Yarullina et al 2014// Appl Biochem Microbiol (Rus)]. Our results indicate that NO, synthesized by Lactobacillus plantarum, takes part in the regulation of intestinal motility in rat [Yarullina et al 2016 // Bull Exp Biol Med]. Involvement of bacterial NO in human cardiovascular system is proposed in the review [Cabrera-Fuentes et al 2016// Basic Res Cardiol].
To study the adhesiveness of Lactobacillus, we evaluate the physicochemical properties of bacterial cell surface [Konnova et al 2013// Chem Commun; Rubtsova et al 2013// Fund Res (Rus); Kirillova et al 2017// Int J Microbiol], and investigate their ability to form biofilms [Yarullina et al 2013// Microbiol (Rus); Bruslik et al 2016// BioNanoSci]. The effects of nitric oxide and iron on biofilm formation by lactobacilli was first shown in our works [Yarullina et al 2013// Microbiol (Rus); Bruslik et al 2016// BioNanoSci].
We also have developed criteria to evaluate Lactobacillus antibiotic resistance on the nutrient agar De Man-Rogosa-Sharpe (MRS). Using this approach, we studied resistance of Lactobacillus strains isolated from fermented dairy products and probiotics, to clinically relevant antibiotics, including those used for eradication of H. pylori, as well as to mesalazine - a drug for the treatment of Crohn's disease and ulcerative colitis [Bruslik et al 2015// Antibiot Khimiot (Rus)]. These results provide reasonable tactical schemes of probiotics application at etiotropic antibiotic therapy.
We also aim to improve the survival of probiotic bacteria in the preparations and in the gastrointestinal tract (GIT). To solve this problem, in collaboration with the Laboratory of Cryochemistry of (Bio)polymers, Institute of Organoelement Compounds (INEOS RAS, Moscow) we suggested to include bacteria into the spatial structure of polysaccharide polymer, such as different types of starch and its individual polysaccharides [Yarullina et al. 2013// Patent RU2491079; Yarullina et al. 2015// Int J Risk Saf Med].
Our work is a key step in developing of effective science-based preparations for medicine and food industry.
Fighting with biofilms on implants: antimicrobial properties of novel derivatives of 2(5H)-furanone
DAAD-0000639905
Project supervisor: I. Sharafutdinov
Biofilms are complex three-dimensional microbial structure often attached to a multitude of surfaces representing the preferred life-style of bacteria in natural and artificial ecosystem. In a biofilm, bacterial cells are embedded in an extracellular matrix of organic polymers such as polysaccharides, peptides and extracellular DNA synthesized and released by the microbes themselves and making bacteria inaccessible for different outer stress factors. The association of an antibiotic with bacterial resistance-modifying agents that increase the activity of the antibiotic has been regarded as a promising strategy. In the context, furanone derivatives might be interesting candidates. Furanones perform many different functions, such as intra- and inter-species signaling and communication, as attractants molecules and pheromones, antimicrobials, and anti-carcinogenes. Several studies have reported the efficiency of synthetic furanones as inhibitors of bacterial biofilms formed by Pseudomonas aeruginosa, Staphylococcus epidermidis, Staphylococcus aureus, Bacillus subtilis, Escherichia coli. Peri-implant infections from bacterial biofilms on artificial surfaces are a common threat to all medical implants. They are a handicap for the patient and can lead to implant failure or even life-threatening complications. New implant surfaces have to be developed to reduce biofilm formation and to improve the long-term prognosis of medical implants and possibility to treat implants with furanones preventing biofouling has to be studied.
The aim of this project is to investigate the antimicrobial activity of 2(5H)-furanone derivatives against biofilm forming oral microflora. A number of 2(5H)-furanone derivatives synthesized in Chemical Institute of Kazan University, among which several compounds exhibited in vitro promise potency to inhibit the biofilm formation by gram positive bacteria such as Staphylococcus
epidermidis, Staphylococcus aureus, Bacillus subtilis will be tested.
13/03/2021