• MeSH
• Biochemistry MeSH
• Metabolic Diseases MeSH
• Pathology MeSH

• Conspectus
• Patologie. Klinická medicína
• Učební osnovy. Vyučovací předměty. Učebnice
• NML Fields
• biochemie
• patologie
• NML Publication type
• učebnice vysokých škol

Guanine quadruplex (GQ) is a noncanonical nucleic acid structure formed by guanine-rich DNA and RNA sequences. Folding of GQs is a complex process, where several aspects remain elusive, despite being important for understanding structure formation and biological functions of GQs. Pulling experiments are a common tool for acquiring insights into the folding landscape of GQs. Herein, we applied a computational pulling strategy─steered molecular dynamics (SMD) simulations─in combination with standard molecular dynamics (MD) simulations to explore the unfolding landscapes of tetrameric parallel GQs. We identified anisotropic properties of elastic conformational changes, unfolding transitions, and GQ mechanical stabilities. Using a special set of structural parameters, we found that the vertical component of pulling force (perpendicular to the average G-quartet plane) plays a significant role in disrupting GQ structures and weakening their mechanical stabilities. We demonstrated that the magnitude of the vertical force component depends on the pulling anchor positions and the number of G-quartets. Typical unfolding transitions for tetrameric parallel GQs involve base unzipping, opening of the G-stem, strand slippage, and rotation to cross-like structures. The unzipping was detected as the first and dominant unfolding event, and it usually started at the 3'-end. Furthermore, results from both SMD and standard MD simulations indicate that partial spiral conformations serve as a transient ensemble during the (un)folding of GQs.

• MeSH
• Biomechanical Phenomena MeSH
• DNA chemistry   MeSH
• G-Quadruplexes * MeSH
• Mechanical Phenomena MeSH
• Molecular Dynamics Simulation * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The enormous sequence heterogeneity of telomerase RNA (TR) subunits has thus far complicated their characterization in a wider phylogenetic range. Our recent finding that land plant TRs are, similarly to known ciliate TRs, transcribed by RNA polymerase III and under the control of the type-3 promoter, allowed us to design a novel strategy to characterize TRs in early diverging Viridiplantae taxa, as well as in ciliates and other Diaphoretickes lineages. Starting with the characterization of the upstream sequence element of the type 3 promoter that is conserved in a number of small nuclear RNAs, and the expected minimum TR template region as search features, we identified candidate TRs in selected Diaphoretickes genomes. Homologous TRs were then used to build covariance models to identify TRs in more distant species. Transcripts of the identified TRs were confirmed by transcriptomic data, RT-PCR and Northern hybridization. A templating role for one of our candidates was validated in Physcomitrium patens. Analysis of secondary structure demonstrated a deep conservation of motifs (pseudoknot and template boundary element) observed in all published TRs. These results elucidate the evolution of the earliest eukaryotic TRs, linking the common origin of TRs across Diaphoretickes, and underlying evolutionary transitions in telomere repeats.

• MeSH
• Transcription, Genetic MeSH
• Nucleic Acid Conformation MeSH
• Evolution, Molecular * MeSH
• Mutation MeSH
• RNA, Plant biosynthesis   chemistry   genetics   MeSH
• RNA Polymerase II metabolism   MeSH
• RNA Polymerase III metabolism   MeSH
• RNA biosynthesis   chemistry   genetics   MeSH
• Sequence Alignment MeSH
• Telomerase biosynthesis   chemistry   genetics   MeSH
• Telomere chemistry   MeSH
• Transcriptome MeSH
• Viridiplantae genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH

Hexameric arginine repressor, ArgR, is the feedback regulator of bacterial L-arginine regulons, and sensor of L-arg that controls transcription of genes for its synthesis and catabolism. Although ArgR function, as well as its secondary, tertiary, and quaternary structures, is essentially the same in E. coli and B. subtilis, the two proteins differ significantly in sequence, including residues implicated in the response to L-arg. Molecular dynamics simulations are used here to evaluate the behavior of intact B. subtilis ArgR with and without L-arg, and are compared with prior MD results for a domain fragment of E. coli ArgR. Relative to its crystal structure, B. subtilis ArgR in absence of L-arg undergoes a large-scale rotational shift of its trimeric subassemblies that is very similar to that observed in the E. coli protein, but the residues driving rotation have distinct secondary and tertiary structural locations, and a key residue that drives rotation in E. coli is missing in B. subtilis. The similarity of trimer rotation despite different driving residues suggests that a rotational shift between trimers is integral to ArgR function. This conclusion is supported by phylogenetic analysis of distant ArgR homologs reported here that indicates at least three major groups characterized by distinct sequence motifs but predicted to undergo a common rotational transition. The dynamic consequences of L-arg binding for transcriptional activation of intact ArgR are evaluated here for the first time in two-microsecond simulations of B. subtilis ArgR. L-arg binding to intact B. subtilis ArgR causes a significant further shift in the angle of rotation between trimers that causes the N-terminal DNA-binding domains lose their interactions with the C-terminal domains, and is likely the first step toward adopting DNA-binding-competent conformations. The results aid interpretation of crystal structures of ArgR and ArgR-DNA complexes.

• MeSH
• Allosteric Regulation MeSH
• Arginine chemistry   metabolism   MeSH
• Bacillus subtilis chemistry   genetics   metabolism   MeSH
• Bacterial Proteins chemistry   genetics   metabolism   MeSH
• Entropy MeSH
• Escherichia coli chemistry   genetics   metabolism   MeSH
• Phylogeny MeSH
• Protein Conformation, alpha-Helical MeSH
• Protein Conformation, beta-Strand MeSH
• Protein Domains MeSH
• Regulon genetics   MeSH
• Repressor Proteins chemistry   genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Sequence Alignment MeSH
• Molecular Dynamics Simulation MeSH
• Protein Binding MeSH
• Hydrogen Bonding MeSH
• Publication type
• Journal Article MeSH

The adverse side effects and acquired resistance associated with the clinical application of traditional platinum-based anticancer drugs have forced investigation of alternative transition metal-based compounds and their cytostatic properties. Over the last years, the anticancer potential of cobalt complexes has been extensively studied, and in-depth analyses of their mode of action have been conducted. In this work, we present antiproliferative activity against human cancer cells of the dinuclear Co(III) complexes bearing the quinizarin ligand and tris(2-aminoethyl)amine (tren, compound 1) or tris(2-pyridylmethyl)amine (tpa, compound 2) co-ligands. To contribute the understanding mechanisms of biological action of these compounds, their association with DNA in the cells, DNA binding in cell-free media, and DNA cleavage capability were investigated in detail. The results demonstrate that both complexes interact with DNA in tumor cells. However, their mechanism of antiproliferative action is different, and this difference is mirrored by distinct antiproliferative activity. The antiproliferative effect of 1 is connected with its ability to intercalate into DNA and subsequently to inhibit activities of DNA processing enzymes. In contrast, the total antiproliferative efficiency of 2, thanks to its redox properties, appears to be connected with its ability to form radicals and, consequently, with the ability of 2 to cleave DNA. Hence, the findings presented in this study may significantly contribute to understanding the antitumor potential of cobalt complexes. Dinuclear Co(III) complexes containing the bioactive quinizarin ligand exhibit antiproliferative activity based on distinct mechanism.

• MeSH
• Anthraquinones chemistry   pharmacology   MeSH
• DNA chemistry   MeSH
• Cobalt chemistry   pharmacology   MeSH
• Coordination Complexes chemical synthesis   chemistry   pharmacology   MeSH
• Humans MeSH
• Ligands MeSH
• Molecular Conformation MeSH
• Tumor Cells, Cultured MeSH
• Cell Proliferation drug effects   MeSH
• Antineoplastic Agents chemical synthesis   chemistry   pharmacology   MeSH
• Drug Screening Assays, Antitumor MeSH
• DNA Cleavage MeSH
• Binding Sites drug effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Circular Dichroic (CD) spectroscopy is one of the most frequently used methods for guanine quadruplex studies and in general for studies of conformational properties of nucleic acids. The reason is its high sensitivity to even slight changes in mutual orientation of absorbing bases of DNA. CD can reveal formation of particular structural DNA arrangements and can be used to search for the conditions stabilizing the structures, to follow the transitions between various structural states, to explore kinetics of their appearance, to determine thermodynamic parameters, and also to detect formation of higher order structures. CD spectroscopy is an important complementary technique to NMR spectroscopy and X-ray diffraction in quadruplex studies due to its sensitivity, easy manipulation of studied samples, and relative inexpensiveness. In this part, we present the protocol for the use of CD spectroscopy in the study of guanine quadruplexes, together with practical advice and cautions about various, particularly interpretation, difficulties.

• MeSH
• Circular Dichroism MeSH
• X-Ray Diffraction MeSH
• DNA chemistry   MeSH
• G-Quadruplexes * MeSH
• Nucleic Acid Conformation MeSH
• Magnetic Resonance Spectroscopy MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

• MeSH
• Allergy and Immunology MeSH
• Immune System MeSH
• Publication type
• Textbook MeSH

• Conspectus
• Patologie. Klinická medicína
• Učební osnovy. Vyučovací předměty. Učebnice
• NML Fields
• alergologie a imunologie

We report frequent losses of components of the classical nonhomologous end joining pathway (C-NHEJ), one of the main eukaryotic tools for end joining repair of DNA double-strand breaks, in several lineages of parasitic protists. Moreover, we have identified a single lineage among trypanosomatid flagellates that has lost Ku70 and Ku80, the core C-NHEJ components, and accumulated numerous insertions in many protein-coding genes. We propose a correlation between these two phenomena and discuss the possible impact of the C-NHEJ loss on genome evolution and transition to the parasitic lifestyle.IMPORTANCE Parasites tend to evolve small and compact genomes, generally endowed with a high mutation rate, compared with those of their free-living relatives. However, the mechanisms by which they achieve these features, independently in unrelated lineages, remain largely unknown. We argue that the loss of the classical nonhomologous end joining pathway components may be one of the crucial steps responsible for characteristic features of parasite genomes.

• MeSH
• Ku Autoantigen chemistry   metabolism   MeSH
• Eukaryota metabolism   MeSH
• Phylogeny MeSH
• Genome MeSH
• Genomics methods   MeSH
• Protein Conformation MeSH
• Models, Molecular MeSH
• DNA End-Joining Repair * MeSH
• Parasites classification   genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Signal Transduction MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Colorectal carcinogenesis is a multistep process involving the accumulation of genetic alterations over time that ultimately leads to disease progression and metastasis. Binding of transcription factors to gene promoter regions alone cannot explain the complex regulation pattern of gene expression during this process. It is the chromatin structure that allows for a high grade of regulatory flexibility for gene expression. Posttranslational modifications on histone proteins such as acetylation, methylation, or phosphorylation determine the accessibility of transcription factors to DNA. DNA methylation, a chemical modification of DNA that modulates chromatin structure and gene transcription acts in concert with these chromatin conformation alterations. Another epigenetic mechanism regulating gene expression is represented by small non-coding RNAs. Only very recently epigenetic alterations have been included in molecular subtype classification of colorectal cancer (CRC). In this chapter, we will provide examples of the different epigenetic players, focus on their role for epithelial-mesenchymal transition and metastatic processes and discuss their prognostic value in CRC.

• MeSH
• Chromatin genetics   metabolism   MeSH
• Epigenesis, Genetic * MeSH
• Epithelial-Mesenchymal Transition MeSH
• Epistasis, Genetic MeSH
• Histones metabolism   MeSH
• Colorectal Neoplasms genetics   metabolism   pathology   MeSH
• Humans MeSH
• DNA Methylation * MeSH
• Cell Transformation, Neoplastic genetics   metabolism   MeSH
• Protein Processing, Post-Translational MeSH
• Disease Progression MeSH
• Gene Expression Regulation, Neoplastic MeSH
• Chromatin Assembly and Disassembly genetics   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

We report the characterization of the dimeric protein AB21 from Agaricus bisporus, one of the most commonly and widely consumed mushrooms in the world. The protein shares no significant sequence similarity with any protein of known function, and it is the first characterized member of its protein family. The coding sequence of the ab21 gene was determined and the protein was expressed in E. coli in a recombinant form. We demonstrated a high thermal and pH stability of AB21 and proved the weak affinity of the protein to divalent ions of some transition metals (nickel, zinc, cadmium, and cobalt). The reported crystallographic structure exhibits an interesting rod-like helical bundle fold with structural similarity to bacterial toxins of the ClyA superfamily. By immunostaining, we demonstrated an abundance of AB21 in the fruiting bodies of A. bisporus.

• MeSH
• Agaricus chemistry   MeSH
• Bacterial Toxins chemistry   MeSH
• Pore Forming Cytotoxic Proteins biosynthesis   chemistry   genetics   MeSH
• Escherichia coli genetics   metabolism   MeSH
• Fungal Proteins biosynthesis   chemistry   genetics   MeSH
• Cations, Divalent chemistry   MeSH
• Protein Conformation MeSH
• Transition Elements chemistry   MeSH
• Recombinant Proteins biosynthesis   chemistry   genetics   MeSH
• Protein Folding MeSH
• Protein Stability MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH