Features of the 8-oxo-7,8-dihydro-2′-dGTP behavior in active site of human DNA polymerase β: structural investigation in silico

The oxidized bases in the composition of DNA as well as DNA precursors (desoxynucleotide triphosphates, dNTPs) appearing in living cell as a result of oxidative stress are the one of major sources of genomic instability. Among oxidized forms of nitrogenous bases, the 8-oxo-7,8-dihydro-2-deoxyguanine (8odG, 8-oxo-dG) is the most ubiquitous. This compound has a high mutagenic potential due to its ability to preferably interact with adenine instead of cytosine. In particular, the 8odG in the composition of the incoming nucleotide triphosphate (8-oxo-GTP) is able to immediately incorporate into the growing DNA chain and, thus, to cause the invert replacement dA → dC because it is possible to pair with the incoming dCTP as well as dATP in the next round of DNA replication. The efficiency of 8oxo-dG incorporation in growing DNA clearly depends on the nature of appropriate DNA polymerases. One of the most sensitive to 8-oxo-dGTP is the eukaryotic DNA polymerase β (pol β). The binding of 8-oxo-dGTP in the active center of pol β can result in two different molecular events. First of them is the incorporation of 8oxoguanine into a growing DNA chain, the other is a discrimination of 8-oxo-dGTP from the active center. While effects of incorporation of this modified guanine in DNA are well studied, the immediate consequences of 8-oxo-dGTP discrimination are still unclear. The behavior of 8-oxo-dGTP molecule in the area of the active site of human DNA polymerase β was investigated using molecular dynamics (MD) calculation. The principle phenomenon revealed as investigation results is existence of two cardinally different models of behavior inherent to 8-oxo-dGTP molecule. In two cases the ligand molecules loses the connections with template dA and starts to migrate inside of enzyme space (migrate trajectories). In the other two cases 8-oxo-dGTP stably stays in DNA polymerase active site, “keeps in touch” with template nucleotide and maintains the hydrogen bonds with it (stable trajectories). The spatial structure of 8-oxo-dGTP in stable trajectories appears to be sufficiently rigid despite the presence of number of bonds around which the free rotation is possible, and its conformational energy is characterized by high stability over the time of studied MD. Average values of energy (-10229.7 and -10227.1 kJ/mol) are practically the same for both cases. Amino acid microenvironment of 8-oxo-dGTP also practically doesn’t change over the studied MD interval. Thus, stable variants of 8-oxo-dGTP behavior evidently correspond to case of the further incorporation modified 8-oxo-dG into growing DNA strand. The behavior of 8-oxo-dGTP molecule in migrate trajectories is significantly more complicated. The 8-oxo-dGTP loses the Н-bonds with template dA6 (at 11 and 6.5 ns of MD in first and second case respectively) and starts to migrate in DNA polymerase space. The 8-oxo-dGTP spatial structure regularly exhibits much more flexibility in comparison to itself behavior in stable trajectories that reflects in corresponded values of individual atomic fluctuations. However, contrary to the expectations the general levels of conformational energy of 8-oxo-dGTP as well as energy fluctuation patterns in both migratory trajectories are completely time stable. The average values of conformational energy are -9938.6 and -10018.6 kJ/mol for trajectories 1 and 2 respectively that is slightly more than corresponded values for stable trajectories. The 8-oxo-dGTP movement pathways of don’t coincide each other that is confirmed by differences of their conformational spaces and amino acid microenvironment. It seems to be the most important that 8-oxo-dGTP not only doesn’t leave the enzyme space but directly prevent transition of DNA polymerase from closed to open conformation as well as the further binding of incoming dNTP. This observation lets a possibility to consider it as natural inhibitor of DNA pol β activity and possible intracellular regulator which mediates the direct transition of the cell from normal state to programmed cell death omitting the malignancy stage

Features of actomyosin superprecipitation reaction in different environment conditions

The influence of divalent cations (Cd2+, Zn2+, Sr2+) and staphylococcus protein A on the superprecipitation reaction of cardiac muscle actomyosin was investigated using methods of preparative protein chemistry, optical spectroscopy and mechanokinetic analysis. It was shown that the metal ions in the range of concentrations 0.1–5 mM inhibit the Mg2+-dependent SPP reaction of actomyosin cardiac muscle and alter the kinetic parameters of this process. It was revealed that protein A modulates superprecipitation dynamics decreasing the maximal value of optical density and time of its half-maximum achievement as well as starting and normalized superprecipitation rates. Thus, investigated factors are able to influence the actin myosin interaction changing their functional parameters of proteins from cardiomyocyte contractile complex. The kinetic characteristics of actomyosin superprecipitation are sensitive to the influence of physicochemical and pharmacological factors and can be used to study their influence on the molecular mechanisms of muscle contraction

Molecular docking of nanosized titanium dioxide material to the extracellular part of GABAB-receptor

A spatial model of nanosized titanium dioxide material was created using Discovery Studio Visualizer software, versions 2.0 and 2.5. A search for and analysis of possible sites of its docking to the extracellular part of GABAB1а receptor subunit were performed using the algorythm for molecular docking PatchDock. The dimensions of the obtained ТіО2 nanoparticle surface were (18.925 × 3.785 × 19.028) Å. Four potentially possible sites of ТіО2 docking to the extracellular part of GABAB1а receptor subunit of GABAB were identified. The ТіО2 nanoparticle demonstrated high affinity of docking to one of the receptor sites with the geometric shape complementarity score of 12562, taking the following values in other sites: 10746; 10370; 10204. The approximate interface area of complex of the extracellular part of GABAB1а receptor subunit of GABAB with ТіО2 for the site with the highest geometric shape complementarity score was 1949.80 Å, and for others – 1273.20 Å, 1261.10 Å and 1170.30 Å, respectively. The evaluation of аtomic contact energy demonstrated the following values for the sites of ТіО2 nanoparticle docking: 362.92; 173.93; 340.63 and 224.61. The nature of connections, stabilizing the sites of ТіО2 docking to the extracellular part of GABAB1а receptor subunit of GABAB, was analyzed in accordance to their amino acid composition

Bioinformatic search of plant microtubule-and cell cycle related serine-threonine protein kinases

A bioinformatic search was carried for plant homologues of human serine-threonine protein kinases involved in regulation of cell division and microtubule protein phosphorylation (SLK, PAK6, PAK7, MARK1, MAST2, TTBK1, TTBK2, AURKA, PLK1, PLK4 and PASK). A number of SLK, MAST2 and AURKA plant homologues were identified. The closest identified homologue of human AURKA kinase was a protein of unknown function, A7PY12/GSVIVT00026259001 from Vitis vinifera (herein named as "STALK", Serine-Threonine Aurora-Like Kinase). Analysis of STALK's three-dimensional structure confirmed its relationship to the subgroup of AURKA-like protein kinases