[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.

[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.

[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.

[Vascular endothelial growth factor and vascular endothelial growth factor receptor 2 in the blood serum, tumor and renal parenchyma of patients with renal cell carcinoma].

Serum, tumor and renal parenchyma levels of VEGF and VEGFR2 were compared in patients with renal carcinoma (RC) with reference to basic clinicomorphological characteristics of the disease. VEGF and VEGFR2 were estimated in 37 RC patients and 57 healthy controls (serum levels only). VEGF and VEGFR2 were detected in all the samples. Their concentrations in the serum were the same in the patients and controls. The tumor tissue contained more VEGF than renal parenchyma. In unfavorable clinicomorphological features the tumor contained higher content of VEGF, higher VEGF/VEGFR2, lower VEGFR2. Thus, angiogenic factors studied closely correlate with clinicomorphological characteristics of renal carcinoma: primary tumor size, stage of the disease, tumor differentiation, tumor pseudocapsule invasion.

[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.

[Left-handed helix of polyproline II type in linker regions of DNA-binding proteins].

Using the databases of proteins and linkers, a search has been undertaken with the aim to find linker segments adopting the polyproline II conformation in DNA-protein complexes. Seventy three linker-DNA complexes were found. It was shown that the mean length of polyproline II segment comprises six residues, praline being not dominant. The symmetry of praline disposition in these regions prevents the formation of the cooperative water net involving amide groups. An example of specific distribution of proline in some proteins of the motility apparatus is presented.

[An analysis of the secondary structure of spider spidroins I and II belonging to different species]. / Analiz vtorichnykh struktur spidroinov pervogo i vtorogo tipov iz paukov, prinadlezhashchikh razlichnym vidam.

We have analyzed the secondary structure of spidroin proteins of I and II types, related to spiders of different species. We used standard methods of secondary structure prediction NNPREDICT and JPRED and also analyzed the occurrences of oligopeptides with a preferred secondary structure with the help of the OLIGON program. We have demonstrated that local segments of the polypeptide chain can adopt alpha- and beta-conformations as well as the left-handed helix of polyproline II type. Periodical patterns found in the amino acid distribution indicate that there is a possibility of development of a macroscopic order accompanied by local conformational transitions.

[A non-statistical approach to the prediction of regular structures in proteins by the example of alpha-helices]. / Nestatisticheskii podkhod k predskazaniiu reguliarnykh struktur v belkakh na primere al'fa-spiralei

A new approach to the analysis of regular structures in proteins that is based on the method of molecular mechanics is proposed. The method uses only the information about the amino acid sequence. The alpha-helical conformation was simulated using the ICM program of molecular mechanics. Energy profiles of the sequences in the alpha-helical conformation, spanning the entire polypeptide chain, were plotted for eight proteins from the Protein Data Bank. The regions of each profile that exhibit energy minima were found to correspond to the alpha-helical regions of the real spatial structure of the protein. Twenty-four out of 25 helices were distinctly pronounced, which indicates a rather high accuracy of the prediction. The energy profiles also help reveal the short regions that correspond to 3/10-helices and the turns that include local alpha-helical conformations. Unlike the known statistical methods of prediction, this method makes it possible to establish the physical principles of the formation of alpha-helical conformations. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2002, vol. 28, no. 6; see also http://www.maik.ru.

[Left-handed helix conformation of poly-L-proline II type in globular proteins. Statistics of incidence and a role of sequence]. / Levospiral'naia konformatsiia tipa poli-L-prolin II v globuliarnykh belkakh. Statistika vstrechaemosti i rol' posledovatel'nosti.

Regions of left-handed polyproline II type conformation in globular proteins were studied throughout the PDB bank. The length and sequence of corresponding fragments were analyzed. It was found that a lot of tetrapeptides (from combinatorial possible ones) show the tendency to be included in the left-handed helices. Much more tetrapeptides do not occur in this structure type.

[A specific complex between a polypeptide in the left helical conformation of polyproline type II and the B-DNA minor groove]. / Spetsificheskii kompleks mezhdu polipeptidom v levospiral'noi konformatsii tipa poliprolin II i oligonukleotidom po maloi borozdke B-formy DNK.

The model of complex formation between polypeptide adopted lefthanded conformation of polyproline II type and minor groove of B-DNA presented. The specific recognition is attained due to possibility of hydrogen bond between Gly NH-group and Thymine O2-group and in addition between Pro CO-group and Guanine NH2-group and between Gly CO-group and Guanine NH2-group. Stereochemical satisfactory of the complex is observed.

[Conformation of a triple collagen spiral as a function of primary structure]. / Konformatsiia troinoi spirali kollagena kak funktsiia pervichnoi struktury.

It has been shown by the method of conformational analysis that conformation of the polypeptide chain of a collagen molecule varies from one type of sequence to another, but in all the sequences the only two types of conformations can be characterized by minimal conformational energy. The collagen molecule as a whole is a combination of the single-bond Rich and Crick model and new double-bonded structure. Rigorous comparison with "thick" conformations obtained by Scheraga and coworkers was undertaken using precise calculations with flexible proline.

[A new approach to the thermodynamic role of imino acids in collagen. Solution of a thermodynamic paradox]. / Novaia traktovka termodinamicheskoi roli iminikislot v kollagene. Razreshenie termodinamicheskogo paradoksa.

The dependence of denaturation transition thermodynamic parameters in various collagens from imino acid compositions has been analysed. Computational and experimental data suggest independence of the collagen molecule hydration on imino acid composition and sequence in the polypeptide chain. The continuous net of hydrogen bonds is interrupted, if imino acid residues occur in the sequence of amino acid residues, as follows from Monte Carlo computations, because the hydrogen of NH-group plays sufficient role in water shell formation for this conformation. As a consequence, entropy of denatured collagen-water system increases hand by hand with increasing imino acid content and therefore delta S increases. The increase of enthalpy of transition from imino acid content is determined by favorable Van der Waals interactions of pyrrolidine rings in native triple helical collagen structure. It was pointed out that proline role is determined by decreasing hydration in the single stranded polypeptide chain in Polyproline II conformation that leads to an increase of entropy of the polypeptide-water system. Thus, the collagen structure formation by imino acids is promoted in the water media due to single chain left-helical conformation being unfavorable for proline residues as well as due to the enthalpy nature of the triple helix stabilization.

[Transitions of the left poly-L-proline II helix-right alpha helix type in the regions of myosin subfragment S2 of cross-striated muscle as a possible conformational basis for a mechanical-chemical act]. / Perekhody tipa levaia spiral' poli-L-prolin II-pravaia alpha-spiral' v uchastkakh subfragmenta S2 miozina poperechnopolosatoi myshtsy kak vozmozhnaia konformatsionnaia osnova mekhano-khimicheskogo akta.

Local conformational states of fibrous fragments of myosin molecules from striated muscle have been studied. Analysis of the amino acid sequences of the rat embryonic skeletal muscle myosin heavy chain and the nematode myosin heavy chain have been performed with the aim to estimate the influence of electrostatic interactions on secondary structure stability of these fragments. The heterogeneity of stability of alpha-helical conformation along the fibrous fragment have been found on the basis of estimation of interaction between side group charges and between side group charges and main chain charges. Periodically located short sections have been found in the N-terminal half of the myosin rod where clusters of Asp and Glu destabilize alpha-helical structure being ionized. Changes of the distribution of charges near the latter sections bring about conformational transitions from left-handed polyproline II helix to right alpha-helix or vice versa. The new scheme of orientation of fibrous part of the cross-bridge in relation to the thick filament for various stages of muscle contraction process suggests asynchronous character of transitions in the different sites. It may be proposed that existence of alteration left- and right-helical fragments in N-terminal half of fibrous part of heavy myosin chain determines zigzag form of this part of myosin molecule in resting muscle. A model of the cross-bridge movement in the course of ATP hydrolysis has been suggested.

[A new type of secondary structure: mobile conformation of polypeptide chain. Data bank for protein structures]. / Novyi tip vtorichnoi struktury: mobil'naia konformatsiia polipeptidnoi tsepi. Analiz banka belkovykh struktur.

As a result of statistical analysis of Protein Data Bank a new type of secondary structure was found in globular proteins. It is mobile (M) conformation, characterised by noncooperative hydration and the increased dynamical properties of the chain. Percentage distribution of amino acid residues between the main secondary structure types is 42.7% for alpha-helix, 19.6% for beta-structure and 19.1% for M-conformation. The most frequently occurring amino acids for M-conformation are proline, cysteine and serine. Fragments of mobile conformation seem to play a major part in local and domain dynamics of protein globule.

[Cyclic conformation of parallel polypeptide chains closed by cross bridges]. / O konformatsiiakh polipeptidnykh parallel'nykh tsepei, zamknutykh v tsikl poperechnymi mostikami.

The method of possible conformations calculations for cyclic structures, formed by two (or more) identical parallel polypeptide chains, closed by cross bridges has been elaborated. The algorithm is necessary for rigorous conformational analysis of cyclic regions in immunoglobulin, fibronectin and myosin.

[Two types of tripeptide conformation in collagen. Calculation of the structure of (Gly-Pro-Ser)n and (Gly-Val-Hyp)n polytripeptides]. / Dva konformatsionnykh tipa tripeptidov v kollagene. Raschet struktury politripeptidov (Gly-Pro-Ser)n i (Gly-Val-Hyp)n.

Conformational analysis of polypeptides (Gly-Pro-Ser)n and (Gly-Val-Hyp)n was carried out for collagen-like triple helical complexes (coiled coils with screw symmetry). The lowest energy structure of the first polymer (helical parameters t 52,8, h 0,282 nm) is very close to that of (Gly-Pro-Hyp)n. The hydroxyl group of a serine residue does not form any intramolecular hydrogen bonds in this structure. (Gly-Val-Hyp)n triple complex is shown to unwind to t 7,7, h 0,297 nm as a result of optimization procedure. These findings confirm the assumption, made earlier on the basis of conformational analysis of (Gly-Pro-Hyp)n, (Gly-Pro-Ala)n, (Gly-Ala-Hyp)n, (Gly-Ala-Ala)n, that the collagen triple helix contains stable wound triplets with proline in the second position, while the absence of imino acid in the 2nd position facilitates the unwinding of the triple helix. Thus, a collagen helix appears to have different parameters for the sites differing in the amino acid sequence. The values measured in the X-ray experiments (h 0,29 nm, t' 36) should be considered as a result of averaging. The model allows to reconcile the X-ray data for collagen and crystalline (Gly-Pro-Pro)10 oligomer.

[Collagen structure with a new principle of forming two nets of inter-peptide hydrogen bonds]. / Struktura kollagena s novym printsipom formirovaniia dvukh setok mezhpeptidnykh vodorodnykh sviazei.

In the new model hydrogen bonds are formed by NH-group of glycine residue in the first polypeptide chain and CO-group of second residue in the tripeptide in the second chain and by NH-group of second residue in the second chain and CO-group of second residue in the first chain etc., i. e. one and the same CO-group serves as acceptor for two NH-groups belonging to other chains. CPK-model was built and then conformational computations for (Gly-Ala-Hyp)n and (Gly-Ala-Ala)n were performed. The resulting optimal structures have unit twist angle t = 52 degrees and 75 degrees. The two-bonded structure may coexist with Rich and Circk type one-bonded structure for any sequence of tripeptides. This suggests the uniform 7/2 helical symmetry for collagen molecule. The model is favourable for hydration and helps to interpret the physico-chemical characteristics of collagen.