Role of structural water for prediction of cation binding sites in apoproteins.

Structures of many metal-binding proteins are often obtained without structural cations in their apoprotein forms. Missing cation coordinates are usually updated from structural templates constructed from many holoprotein structures. Such templates usually do not include structural water, the important contributor to the ion binding energy. Structural templates are also inconvenient for taking into account structural modifications around the binding site at apo-/holo- transitions. An approach based upon statistical potentials readily takes into account structural modifications associated with binding as well as contribution of structural water molecules. Here, we construct a set of statistical potentials for Mg2+, Ca2+, and Zn2+ contacting with protein atoms of a different type or structural water oxygens. Each type of the cations tends to form tight contacts with protein atoms of specific types. Structural water contributes relatively more into the binding pseudo-energy of Mg2+ and Ca2+ than of Zn2+. We have developed PIONCA (Protein-Ion Calculator), a fast CUDA GPGPU-based algorithm that predicts ion-binding sites in apoproteins. Comparative tests demonstrate that PIONCA outperforms most of the tools based on structural templates or docking. Our software can be also used for locating bound cations in holoprotein structures with missing cation heteroatoms. PIONCA is equipped with an interactive web interface based upon JSmol.

[Structural and Physicochemical Characteristics of Conformationally Stable alpha-Helical Oligopeptides].

The analysis of conformationally stable (conformational conservative) tetrapeptides selected from protein structures deposited in PDBSelect data bank has been fulfilled. The subset contained 943 tetrapeptide amino acid sequences and there were merely five 3D protein segment representatives for each sequence. As a result, the conclusion has been drawn on the basis of DSSP annotation analysis that in the majority of cases (900 of 943) alpha-helical conformation is obvious. Different than alpha-helix, in particular, the left-handed polyproline II helical conformation was observed in 43 sequences. The physical and chemical properties of conformationally stable peptides taken from the appropriate sample were estimated by the average hydrophobicity/hydrophilicity of tetrapeptides. The results of calculations show that the "neutrality" towards hydrophobicity/hydrophilicity is representative of conformationally stable oligopeptides. It should be noted, that dispersion of hydrophobicity/hydrophilicity distribution is sufficiently lower than for the test subsets. Thus, the conformationally stable oligopeptides present a distinct group of local protein structures which are very close with respect to conformational and physicochemical properties. In accordance with our developed theory of specific long range interactions these peptides are the objects being quite useful for effective mutual molecular recognition.

[Disallowed conformations of polypeptide chain exemplified by the ß-bend of the ß-hairpin in the α-spectrin CH3-domain].

The work presents the results of an exhaustive conformational analysis of ß-turns involving amino acid residues with disallowed backbone conformation of the polypeptide chain. It is known that the first residue of the ß-turn (Asn47) of the distal ß-hairpin in the α-spectrin SH3-domain is characterized by sterically disallowed main chain conformation (values of the dihedral angles (φ and ψ are in the right bottom quadrant of the Ramachandran plot). All α-spectrin structures with the anomalous elements deposited in the PDB were analysed. We hypothesized that the formation of disallowed conformation may occur through the fixation (due to the SH3 domain structure) of the adjacent to the ß-turn amino acid residues with the ß-structure. These residues are disposed in such a manner that ß-turn conformation of the residues contributes just to the disallowed local conformation of this residue whereas any other ß-turn conformations (with allowed local conformation) are impossible. To test this hypothesis an exhaustive conformational analysis of the ß-bend has been performed by altering internal coordinates (two pairs of φ and ψ angles and two Ω angles). The conformations were selected as a result of grid search procedure with. 1 degrees step that corresponded to stereochemically allowed local deformations of the polypeptide chain segment forming the ß-turn. In all conformations obtained the local conformation of Asn47 rests in the disallowed region. The conformations found include conformations coinciding with experimentally determined structures from the PDB as well as an additional variant that differs from X-ray structure in values of a pair of φ and ψ angles of the second residue belonging to the ß-bend. Values of these angles fall in the region of the Ramachandran plot near the line φ = 0 (and negative values of ψ) i.e. in strongly disallowed region without experimental points. Therefore the additional variants of the ß-turn local deformation are impossible to observe in experiment. Thus, the idea that disallowed conformation is intruded to the ß-bend by fixation of adjacent residues receives confirmation in this work. The topological limitations in a context of the structure in such kind of ß-hairpins exclude the allowed local conformations.

["Co-phased blocks"--conservative regions of double-stranded DNA of transcriptional regulatory modules--are close in space due to phasing at nucleosome DNA super helix].

Conservative regions varying in length from 30 to 70 nucleotides (nt) are revealed as a result of interspecies comparison of homologous nucleotide sequences of the loci, genes controlling early development in Drosophila species. These regions are observed at the intermediate level between transcription factor binding sites (usually about 7-10 nt) and the nucleosome repeat unit (165-210 nt). Segments found are located mainly in the area of known functional elements of the locus (cis-regulatory modules: enhancers, a proximal promoter, a coding segment). Conservative domains occupy less than a half of the full enhancer length and contain practically all annotated transcription factor binding sites. A quasi periodical pattern of conservative region disposition is in accord with the experimental nucleosome localization pattern. The distance between neighboring domains is about 84 nt, i.e. a pitch of nucleosome DNA super helix. A repeatable accuracy of the distance permits of treating the delineated conservative regions as co-phased blocks and stating that these domains are close in the space being located at neighboring coils of the nucleosome DNA super helix.

[On efficient in vitro purification of cell suspensions containing malignant cells].

We propose a method for magnetic sorting of cell suspensions able to differentiate not only cells with single specific antigen on their surface, but also cells with a group of pre-defined antigens. Individually, each antigen of this group may be present on surfaces of non-selected cells. However, only the simultaneous presence of all given antigens on the cell surface means that such a cell should be separated. The method is of interest, for purification of cell suspensions from malignant cells, in particular, for purification of bone marrow material for autologous transplantation in leukemia.

[Classification of amino acids based on a comparative analysis of contacts in DNA-protein complexes and specific DNA-protein interactions].

The classification of amino acid residues based on the events of contact formation between distinct amino acid and selected nucleotides was constructed. Thus, the most integral properties, that characterize interactions in organization of DNA-protein complexes, were used. We applied the Voronoi-Delaunay tessellation to draw statistics of contacts and area of contacts for the set included 1937 DNA-protein complexes. Similarities of amino acid residues have been searched for based on the comparison of corresponded rows and matrixes of contacts and areas of contacts. Nine measures of distance were used for estimation of rows similarity degree. The procedure of clustering amino acids in groups included three hierarchical and two nonhierarchical methods. A total tree was built using nine techniques of estimating distance with three hierarchical clustering methods. It was shown that clustering centers in the main groups are always constant while other relationships between objects vary. Clustering of binary associations was found for the most amino acids. Major classes of up to six amino acids correspond to the certain local structures of the polypeptide chain in the context of amino acid composition. These data should be taken into account when designing DNA-protein ligands.

[Conformationally stable segments in helical structures of polypeptide chains of proteins and their role in high level structures formation].

In the work the arguments are presented in favor of the idea on the role of conformationally stable oligo-peptides in specific long-distance interactions in phenomena of molecular recognition during various biological processes. Original authors' and literature data are taken into account. The examples of conformationally stable short oligopeptides participation in alpha-helix and collagen type structures formation are given simultaneously with theoretical approaches. The conformationally stable oligopeptides obtained in the course of PDB bank analysis are discussed. The role of amino acids sequence in collagen helix formation is shown.

[Mutual disposition of nucleosomes and exons differs from common genome pattern at DNA segments containing periodic nucleotide sequences].

The correlation was observed between DNA segments characterized by periodic patterns of nucleotides and nucleosome DNA binding sites predicted by the profiles of local DNA flexibility. Exons of human type I and VII collagen genes loci have been chosen as a sample set of periodic DNA segments. It was pointed that short periodic exons coding fibrillar part of these proteins are localized near borders of the DNA sequence that binds the histone core. One of the borders of exon and of this sequence coincide as a rule. In non-fibrillar parts of collagen genes exons (longer on average) dispose, in contrast, in any place of the DNA sequence bound to the nucleosome core.

[DNA sequencing using specific long-range interaction between macromolecules].

On the basis of the theory of specific long-range interaction between long molecules the approach has been elaborated for the "fast reading" of nucleotide sequences in sole DNA molecule. Firstly, a stretching force applies to the molecule that causes its unwinding from B-form to so called S-form. Then, a molecule disposes in a stretched state on the background. After this, the electrostatic potential is estimated in the space along the DNA filament. Information obtained is sufficient for the deduction of the nucleotide sequence in DNA. Another approach to the "reading" of information reduces to the measurement of the filament element deformation induced by electrostatic field from the electrode that stretches the filament by alternating current applied.

[Interconnection between architecture of protein globule and disposition of conformational conservative oligopeptides in proteins from one protein family].

It was shown that selective interactions between helical segments of macromolecules can realize in globular proteins in the segments characterized by the same periodicities of charge distribution i.e. between conformationally conservative oligopeptides. It was found that in the macromolecules of alpha-helical proteins conformationally conservative oligopeptides are disposed at a distance being characteristic of direct interactions. For representatives of many structural families of alpha-type proteins specific disposition of conformationally conservative segments is observed. This disposition is inherent to a particular structural family. Disposition of conformationally conservative segments is not related to homology of the amino acid sequence but reflects peculiarities of native 3D-architectures of protein globules.

[On the optimal folding of protein molecules].

The process of globular structure formation from a long molecular chain has been examined. In the course of this process, various regions of the chain interact with one another. We classify the bonds formed during this process as "correct" and "erroneous" ones. The term "correct" bonds implies the bonds characteristic for a completely formed native globular structure. All other bonds can be treated as "erroneous". It was demonstrated that the process of globule formation may proceed actually without the formation and the following decay of "erroneous" bonds. Our model permits one to avoid the examination of numerous "erroneous" variants since, between the regions of the chain that form "correct" bonds, long-distance interactions characterized simultaneously by high selectivity take place. The existence of interactions of this kind facilitates the drawing together and subsequent interaction of just these regions of the chain that yield "correct" bonds. Based on the data bank analysis, it was demonstrated that the model elaborated is valid not only for abstract structures but also for real polypeptide chains capable of forming protein globules and superhelical fibrils.

A deeper look into transcription regulatory code by preferred pair distance templates for transcription factor binding sites.

MOTIVATION: Modern experimental methods provide substantial information on protein-DNA recognition. Studying arrangements of transcription factor binding sites (TFBSs) of interacting transcription factors (TFs) advances understanding of the transcription regulatory code. RESULTS: We constructed binding motifs for TFs forming a complex with HIF-1α at the erythropoietin 3(')-enhancer. Corresponding TFBSs were predicted in the segments around transcription start sites (TSSs) of all human genes. Using the genome-wide set of regulatory regions, we observed several strongly preferred distances between hypoxia-responsive element (HRE) and binding sites of a particular cofactor protein. The set of preferred distances was called as a preferred pair distance template (PPDT). PPDT dramatically depended on the TF and orientation of its binding sites relative to HRE. PPDT evaluated from the genome-wide set of regulatory sequences was used to detect significant PPDT-consistent binding site pairs in regulatory regions of hypoxia-responsive genes. We believe PPDT can help to reveal the layout of eukaryotic regulatory segments. CONTACT: ivan.kulakovskiy@gmail.com SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The intramolecular impact to the sequence specificity of B-->A transition: low energy conformational variations in AA/TT and GG/CC steps.

It is well known, that local B--> A transformation in DNA is involved in several biological processes. In vitro B<--> A transition is sequence-specific. The physical basis of this specificity is not known yet. Here we analyze the effect of intramolecular interactions on the structural behavior of the GG/CC and AA/TT steps. These steps exemplify sequence specific bias to the B- or A-form structure. Optimization of potential energy of the molecular systems composed of an octanucleotide, neutralized by Na(+) and solvated with TIP3P water molecules in rectangular box with periodic boundary conditions gives the statistically representative sets of low energy structures for GG/CC and AA/TT steps in the middle of the diverse flanking sequences. Permissible 3D variations of GG/CC and AA/TT, and correlation of the relative motion of base pairs in these steps were analyzed. AA/TT step permits high variability for low energy conformers in the B-form DNA and small variability for low energy conformers in the A-form DNA. In contrast GG/CC step permits high variability for low energy conformers in the A-form DNA and small variability for low energy conformers in the B-form DNA. The relative motion of base pairs in GG/CC step is high correlated, while in AA/TT step this correlation is notably less. Atom-atom interactions inside-the-step always favors the B-form and their component - stacking interactions (atom-atom interactions between nucleic bases) is crucial for the duplex stabilization. Formation of the A-form for both steps is a result of interactions with the flanking sequences and water-cation environment in the box. The average energy difference between conformations presenting B-form and A-form for the GG/CC step is high, while for the AA/TT step it is rather low. Thus, intramolecular interactions in GG/CC and AA/TT steps affect the possible conformational diversity ("conformational entropy") of the A- and B- type structures of DNA step. This determines the known bias of the A-form DNA depending on the enrichment of sequences with GG/CC. If structural tuning during the process of protein-DNA complex formation lead to the local B--> A transformation of DNA, it is largely directed by high conformational diversity of GG/CC step in the A-form. In such a case the presence in the target site of both kinds of examined steps ensures the reversible character of ligand binding.

The Intramolecular Impact to the Sequence Specificity of B→A Transition: Low Energy Conformational Variations in AA/TT and GG/CC Steps.

Abstract It is well known, that local B→A transformation in DNA is involved in several biological processes. In vitro B↔A transition is sequence-specific. The physical basis of this specificity is not known yet. Here we analyze the effect of intramolecular interactions on the structural behavior of the GG/CC and AA/TT steps. These steps exemplify sequence specific bias to the B- or A-form structure. Optimization of potential energy of the molecular systems composed of an octanucle-otide, neutralized by Na(+) and solvated with TIP3P water molecules in rectangular box with periodic boundary conditions gives the statistically representative sets of low energy structures for GG/CC and AA/TT steps in the middle of the diverse flanking sequences. Permissible 3D variations of GG/CC and AA/TT, and correlation of the relative motion of base pairs in these steps were analyzed. AA/TT step permits high variability for low energy conformers in the B-form DNA and small variability for low energy conformers in the A-form DNA. In contrast GG/CC step permits high variability for low energy conformers in the A-form DNA and small variability for low energy conformers in the B-form DNA. The relative motion of base pairs in GG/CC step is high correlated, while in AA/TT step this correlation is notably less. Atom-atom interactions inside-the-step always favors the B-form and their component - stacking interactions (atomatom interactions between nucleic bases) is crucial for the duplex stabilization. Formation of the A-form for both steps is a result of interactions with the flanking sequences and water-cation environment in the box. The average energy difference between conformations presenting B-form and A-form for the GG/CC step is high, while for the AA/TT step it is rather low. Thus, intramolecular interactions in GG/CC and AA/TT steps affect the possible conformational diversity ("conformational entropy") of the A- and B- type structures of DNA step. This determines the known bias of the A-form DNA depending on the enrichment of sequences with GG/CC. If structural tuning during the process of protein-DNA complex formation lead to the local B→A transformation of DNA, it is largely directed by high conformational diversity of GG/CC step in the A-form. In such a case the presence in the target site of both kinds of examined steps ensures the reversible character of ligand binding.

[Periodicities in nucleotide distribution in the locus of the light chain of Gallus gallus immunoglobulins].

Fourier spectra of nucleotide sequences from chromosome 15 of Gallus gallus have been obtained. The periodical structures of several Ig locus fragments have been characterized. The uniqueness of Fourier spectra of the locus of the Ig light chain and its fragments (such as a variable region, a constant region, an enhancer, and several other fragments) among Fourier spectra of other fragments of chromosome 15 is demonstrated.

[Left helix of polyproline II type and genesis of beta-structures in spidroins 1 and 2 and their recombinant analogs].

The distribution of secondary structures along the polypeptide chains of spider proteins spidroins 1 and 2 and their recombinant analogs has been studied by statistical methods. It was found that these proteins in the monomolecular form contain only trace amounts of beta-structures. At the same time, the regions of the sequence including Ala and Gly are predicted as helical-containing (with alpha-helices and left-helices of polyproline II type). An analysis of literature data and our data obtained in this study shows that the main conformation of the polypeptide chain solutions of spidroins 1 and 2 and their recombinant analogs in water solutions is the left-helix of polyproline II type with some contaminations of alpha-helices and a very small share of beta-structures. The transition to the state with extended conformations, which are peculiar to mature filaments of spider webs, requires the dehydration of the polypeptide chain backbone. Thus, the genesis of beta-structure in spider web proteins is determined by the conditions of conformation transitions between the main regular conformations of the polypeptide chain backbone.

[Mutual disposition of short conformationally stable oligopeptides in the 3D structure of globular proteins].

The disposition of conformationally stable tetrapeptides and the segments containing these peptides in polypeptide chains of proteins from different structural and functional groups has been analyzed. It was shown that the disposition of segments of this kind can be treated as a statistical one. More than 60% of stable peptides in the globula space are separated by distances no greater than 5 A.

[On the mechanisms of the influence of imino acids on the physical characteristics of collagens].

The influence of imino acids on the thermodynamic characteristics of collagen type structures in various collagens has been analyzed. It was shown that the basic mechanism of entropy increase in the protein-water system consists in the alteration in the number of cooperative segments accompanying the increase in imino acids content, which can be observed during the melting of the fibrous macromolecule. The range of variation in the physical characteristics of cooperative units is determined, in particular, by the variability of hydrogen bond parameters. This is displayed in a broadening of the bands of NH-valence vibrations and the half-widths of transitions in post-denatured structures. Thus, the basic mechanism of the influence of imino acids on thermodynamic characteristics of collagens is related to the complex nature of the melting process. The dehydration-hydration mechanisms of native and denatured states become significantly different upon replacement of any amino acid by an imino acid.

[Geometrical analysis of protein-DNA interactions on the basis of the Voronoi-Delaune tessellation].

A new method for the rigorous analysis of DNA-protein contacts has been developed on the basis of Voronoi tessellation. This method permits one to determine close neighbors on the atomic level and compute the area of contact by edges of Voronoi polyhedra. Based on the results of the study of 1109 protein-DNA complexes from PDB, it was demonstrated that about one third of the contacts are the contacts with positively charged Arg and Lys. There is distinct amino acid prevalence for nucleotides: for A - Pro; for T - His; for G - Asp; for C - Trp, Asp, Glu. Therefore, GC pairs prefer to interact with negatively charged residues, and alanine and methionine, whereas AT pairs prefer to interact with histidine and unpolar residues.

[Nucleosomal repeat and the location of exons and introns in genes of collagens types I and VII].

Regions with a quasiperiodical location of exon--intron sites have been found in the loci of genes of I and VII type collagens (with a total length of exons more than 15% of the entire size of the locus). The periods observed are similar to periods typical for the nucleosomal level of the organization of chromatin. It was shown that the sites consisting of successively arranged exons and introns form groups involving two to five such regions of the same length. The groups encoding the fibrillar regions of the gene product contain more than 50% of exons. The regions are on the average 165 nt long, which is close to the minimal nucleosomal repeat length observed in some regions of the eukaryotic genome. In the nonfibrillar region of the gene of VII type collagen, groups of several exon-intron pairs with an average length of 227 nt were identified. The change in the length of exon-introns sites on going from the nonfibrillar to the fibrillar moiety occurs in a jump, which is clearly seen on a periodogram of the locus.