[Spatial structure of the rp142 peptide containing the immunodominant epitope of the HIV-1 gp120 protein. A theoretical study]. / Prostranstvennaia struktura peptida rp142, soderzhashchego immunodominantnyi épitop belka gp120 VICh-1. Teoreticheskoe issledovanie.

A model of spatial structure of the synthetic peptide rp142 (24 amino acid residues) containing the immunodominant epitope of the HIV-1 protein gp120 in the region Gly-10-Phe-15 was constructed by the method of "constrained" molecular mechanics, which uses the algorithms of theoretical conformational analysis, based on NMR spectroscopy data. A comparative analysis of calculated conformations revealed that the spatial structure of rp142 in solution can be described by a family of conformations to which nine different structural clusters involving the sets of topologically close conformers correspond. It is shown that the main chain of the peptide forms irregular but "structured" conformations in which the main portion of amino acid residues is incorporated into beta-turns and helix-like fragments, while Pro-11 and Gly-12 form in some structures inverse gamma-turns, which rarely occur in protein-peptide molecules. It was found that the spatial packing of the Gly-10-Phe-15 hexapeptide in different clusters is realized at different internal rotation angles, to which topologically close structures correspond. It is assumed that this invariant structural element describes the "conformation of complex formation" that is complementary to the antigen-binding center of antibodies and is responsible for their binding to the peptide.

Melting of cross-linked DNA. III. Calculation of differential melting curves.

In our previous papers I and II (D. Y. Lando et al, J. Biomol. Struct. Dynam. (1997) v. 15, N1, p. 129-140, p. 141-150), two methods were developed for calculation of melting curves of cross-linked DNA. One of them is based on Poland's and another on the Fixman-Freire approach. In the present communication, III, a new theoretical method is developed for computation of differential melting curves of DNAs cross-linked by anticancer drugs and their inactive analogs. As Poland's approach, the method allows study of the influence of the loop entropy factor, delta(n), on melting behavior (n is the length of a loop in base pairs). However the method is much faster and requires computer time that inherent for the most rapid Fixman-Freire calculation approach. In contrast to the computation procedures described before in communications I and II, the method is suitable for computation of differential melting curves in the case of long DNA chains, arbitrary loop entropy factors of melted regions and arbitrary degree of cross-linking including very low values that occur in vivo after administration of antitumor drugs. The method is also appropriate for DNAs without cross-links. The results of calculation demonstrate that even very low degree of cross-linking alters the DNA differential melting curve. Cross-linking also markedly strengthens the influence of particular function delta(n) upon melting behavior.

Melting of cross-linked DNA: II. Influence of interstrand linking on DNA stability.

In the previous paper (D.Y. Lando, J. Biomol. Struct. Dynam, 15, 129-140 (1997)) the melting of cross-linked DNA with N base pairs and omega interstrand cross-links has been considered theoretically. In the present study on the basis of these results, two simple schemes are developed for the computation of melting curves of cross-linked DNA. The investigation of influence of interstrand linking on DNA stability has been carried out by computer simulation. It is shown that the relative concentration of cross-links, CCT = omega/N, their distribution along a DNA molecule, and particular values of the entropy factors of small loops formed by cross-links in melted regions strongly affect the DNA melting temperature, Tm. On the contrary, for DNA without cross-links, a ten-fold increase or decrease in the entropy factors of small loops does not cause the Tm variation. The comparison of the results of calculation with experimental data suggests that the majority of types of cross-link neither maintain ordered parallel orientation of bases in melted regions nor increase considerably the thermostability of cross-linked base pairs. Four different ways of influence of interstrand cross-linking on the DNA double helix stability are considered. It is shown that cross-linking significantly enhances the influence of single strand stiffness in melted regions on DNA melting behavior.

[Synthesis and properties of membrane-addressed antioxidants based on phosphorylated pyrocatechols]. / Sintez i svoistva membranoadresovannykh antioksidantov na osnove fosforilirovannykh pirokatekhinov.

Amphiphilic membrane-addressed antioxidants differing in their hydrophobic-hydrophilic balance, 3,5-di-tert-butyl-2-hydroxyphenyl phosphate, (3,5-di-tert-butyl-2-hydroxyphenyl)hexadecyl phosphate, and (3,5-di-tert-butyl-2-hydroxyphenyl)-5-cholesten-3 beta-yl phosphate, were synthesized starting from 3,5-di-tert-butyl-o-quinone. Even at 10(-8) M concentration, 3,5-di-tert-butyl-2-hydroxyphenyl phosphate and (3,5-di-tert-butyl-2-hydroxyphenyl)hexadecyl phosphate inhibit formation of malonic aldehyde during peroxidation of lipids from rat brain homogenate initiated with a Fe(2+)-ascorbate system. At 10(-4) M and higher concentrations of antioxidants in the brain homogenate, the formation of malonic aldehyde is inhibited more efficiently than with tocopherol at the same concentrations.

Theoretical and experimental study of DNA helix-coil transition in acidic and alkaline medium.

The theoretical approach to the calculation of the influence of selective binding of small ligands on DNA helix-coil transition has been described in the previous paper (Lando D. Yu., J. Biomol. Struct. Dyn., (1994)). In the present paper that method is used for the study of DNA protonation and deprotonation in acidic and alkaline medium by theoretical analysis of pH effect on DNA heat denaturation. The mechanism of DNA protonation in acidic medium and pK values of nucleotides are well known. It gave us an opportunity to check the theory without any fitting of pK values. A good agreement between experimental and calculated functions Tm(pH) and delta T(pH) (melting temperature and melting range width) obtained for acidic medium proved the validity of the theory. However, for alkaline medium there was not even qualitative agreement when the agreed-upon mechanism of deprotonation was considered. Looking into the cause of the discrepancy, we have studied the DNA melting for different mechanisms of deprotonation by calculation of Tm(pH) and delta T(pH). As a result, it has been established that the discrepancy is due to deprotonation of bonded GC base pairs of helical DNA regions (pK = 11). It was shown that the early known protonation and newly found deprotonation of helical DNA essentially stabilised double helix in alkaline and acidic medium.

[Model of the spatial structure of peptide T]. / Model' prostranstvennoi struktury peptida T.

The spatial structure model of peptide T (AIDS reproduction inhibitor, the amino acid sequence of which corresponds to the fragment into the binding site of the virus protein gp120 with T4 receptor) is proposed. Peptide structure modelling has been carried out by the previously developed method based on joint usage of the molecular mechanics algorithms and NMR spectroscopy data. To build the model, two-dimensional nuclear Overhauser effect spectroscopy data for RNase A homologous fragment 22-26 were taken from the literature. The result of the presented work was a set consisting of six types of low-energy structures with different spatial packing of the peptide main chain. All structural types have been shown to be characterized by the lack of strict determination of the side chain conformations of the amino acid residues that can be realized in a few states providing approximately equal (within the given type) stabilization of one main chain form. At the same time, despite the definite differences, all of the selected structures were characterized by the presence of two consecutive reverse polypeptide chain turns at the C-terminal pentapeptide fragment. This site is supposed to be responsible for the peptide binding with T4 receptor and the antiviral effect.

[The resistance of liposomes as potential drug carriers to the action of phospholipase A2]. / Izuchenie ustoichivosti liposom kak potentsial'nykh perenoschikov lekarstvennykh veshchestv k deistviiu fosfolipazy A2.

Stability of liposomes with varied phospholipid (PL) content towards the action of phospholipase A2 (PLA2) was studied in vitro. Liposomes were prepared of phosphatidylcholine (PC), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), and phosphatidylinositol (PI) and their mixtures with sphingomyelin (SM). PC and PG liposomes were also treated with PLA2 in the presence of a number of proteins such as cytochrome c, cytochrome b5, cytochrome P-450, polylysine, and histone H1. Among all the liposomes studied PG-containing ones were found to be most stable towards PLA2. In SM-containing liposomes, the levels of PG, PI, PE and PC after 20 min of hydrolysis were 65, 62, 55 and 50%, respectively. In PC + PG liposomes, the PC content after treatment with PLA2 was by about 15-20% higher than in control PC liposomes. The addition of all the proteins studied influenced significantly stability of both PC and PG in two-component PL liposomes, but had no effect on liposomes consisting only of PC. The incorporation of the integral membrane protein cytochrome P-450 into PC and PG liposomes caused a decrease in their stability towards PLA2, the decrease depending on the lipid/protein molar ratio.

[Lipolysis in model membranes in the presence of positively charged soluble proteins]. / Issledovanie lipoliza v model'nykh membranakh v prisutstvii polozhitel'no zariazhennykh rastvorimykh belkov.

Phospholipase A2 hydrolysis of neutral and negatively charged lipid membranes modified by positively charged proteins has been studied using liposomes composed of either dioleoylphosphatidylcholine (DOPC) or dioleoylphosphatidylglycerol (DOPG) alone or their equimolar mixture in the presence of cytochrome c, histone H1, cytochrome b5, and polylysine. Twenty minutes after the reaction had been initiated, DOPC hydrolysis was 58%, while that in the equimolar mixture with DOPG was 35%. DOPG hydrolysis was more complete in binary mixtures of liposomes. The same was observed for liposomes in the presence of cytochrome c. Hydrolysis of phospholipids in binary liposomes in the presence of histone H1 was 3 times faster than that in protein-free liposomes. In the presence of polylysine the rate of DOPG hydrolysis was decreased. The results obtained are suggestive of electrostatic interactions between hydrophilic proteins and negatively charged phospholipids, the phospholipase A2 catalytic activity being affected by these interactions.

[Spatial structure of apamin in solution]. / Prostranstvennaia struktura apamina v rastvore.

The calculation of the complete spatial structure of the bee venom peptide neurotoxin apamin has been carried out by means of a method elaborated earlier. It is based on the joint utilization of the molecular mechanics algorithms and NMR spectroscopy data. It was established that the molecule backbone conformation in solution may be represented as the combination of the beta-turn III (residues 2-5) and alpha-helical segment (9-18) both separated by the non-standard bend IV (5-8). The most probable system of the intramolecular hydrogen bonds in the apamin polypeptide backbone was proposed. Certain amino acid residues have been shown to be characterized by the lack of strict determination of the conformations of their side chains which may be realized in a few states providing approximately equal stabilization of the same form of the main chain. The conformational parameters of the proposed apamin structural model are appropriate to the NMR spectroscopy data derived from the literature and used in the calculations and are not contradictory to other experimental information.

[Local structure of cytochrome c from horse heart in solution. Conformational analysis using data of two-dimensional nuclear Overhauser effect spectroscopy]. / Lokal'naia struktura tsitokhroma c iz serdtsa loshadi v rastvore. Konformatsionnyi analiz po dannym dvumernoi spektroskopii iadernogo éffekta overkhauzera.

Using the earlier suggested method the calculation of the backbone conformations of horse heart cytochrome c in oxidized (ferricytochrome c) and reduced (ferrocytochrome c) states has been performed by the two-dimensional nuclear Overhauser effect spectroscopy data. For both protein forms the secondary structure elements have been revealed and the conformations of the irregular polypeptide chain segments have been analysed. The similarity of the secondary structures of ferri- and ferrocytochrome c in solution was established from the comparison of their conformations. Small differences between the conformations of two molecule forms are shown to be localized within the polypeptide chain fragments situated in the spatial structure near the heme crevice. The comparison of the dihedral phi and psi angles in the calculated conformations of horse cytochrome C with the corresponding characteristics of X-ray structures of tuna ferri- and ferrocytochrome c made for the oxidized and reduced protein forms using the quantitative criteria testifies the similarity of their conformations in solution and crystal. In is shown that the conformational changes of the separate amino acid residues which take place as the result of the "solution-to-crystal" transition occur on the surface fragments of protein globule and do not lead to essential alterations of the secondary molecule structure.

A method for determining the relative effect of ligands on A-T and G-C base pairs in DNA: applications to metal ions, protons and two amino acids.

A new method is described for the study of specific interactions of low-molecular ligands with the base pairs of DNA. This method is based on the comparative analysis of melting temperature changes in DNAs of different GC-content in the presence of low molecular weight ligands. In this paper, the method is applied to Mn2+, Ni2+, Co2+ ions, deprotonation, amino acids, beta-alanine and gamma-aminobutyric acid (gamma-ABA). Differences in Tm are affected not only by the changes of relative stability of AT- and GC-pairs, but also by other factors. A theoretical analysis of the sequence specificity of low-molecular ligands on the base pairs in DNA molecules characterized by a high degree of sequence heterogeneity is also presented.

[Regulation of the catalytic activity of the monooxygenase enzyme system depending of the substrate structure and phospholipid composition of the model membrane]. / Reguliatsiia kataliticheskoi aktivnosti monooksigenaznoi fermentoi sistemy v zavisimosti ot struktury substrata i fosfolipidnogo sostava model'noi membrany.

A comparative study of pentoxy- and benzyloxyresorufin dealkylation by a monooxygenase enzyme system in dilauroylphosphatidylcholine micelles and in proteoliposomes was carried out. In proteoliposomes whose lipid matrix is formed by double phospholipid mixtures, a cooperative regulation of the oxidation reaction was found. It was shown that the cooperativity of this process depends on the substrate type as well as on the phospholipid composition of the vesicles and decreases during peroxide oxidation of lipids. The results obtained are discussed in terms of protein-protein and protein-phospholipid interactions in the oligomer ensembles consisting of several cytochrome P-450 LM2 molecules.

[The effect of long-range interactions between adsorbed ligands on the DNA helix-coil transition]. / Vliianie vzaimodeistviia dal'nego poriadka mezhdu adsorbirovannymi ligandami na perekhod spiral'-klubok DNK.

The influence of different types of long-range interaction of ligands adsorbed on DNA on the helix-coil transition was theoretically considered. The contact interaction was shown to differ significantly from the long-rang one. It was shown also that even weak dependence of a long-range potential on a degree of helicity resulted in the strong changes of a DNA melting curve. This result allowed to understand the different experimental data on DNA melting in the presence of different substances which reduced AT-and GC-base pairs thermostability difference.