Structure-functional organization of the secretin molecule. II. Direct conformation problem

Spatial structure of peptide hormone secretin was investigated by the theoretical conformational method. A solution of the "direct conformational problem" for this hormone indicated that the possible structure of the secretin molecule under polar conditions may be described only by two families of low-energy conformations, possessing relatively conformational valid (Thr7-Leu22) and variable (His1-Phe6 and Leu23-Val27NH2) fragments. One of these families is comprised by five twists of the alpha-helix, while the second isoenergetic family possesses two short segments of the alpha-helix, divided by an irregular structure of the tetrapeptide.     

The structure-function organization of neurokinin A and neurokinin B molecules. II. Theoretical conformational analysis of neurokinin B

The spatial structure of a neurokinin B molecule was investigated by the method of theoretical conformational analysis. The conformational analysis of this molecule indicated that the possible structure of neurokinin B under polar conditions may be described by five families of low-energy conformations possessing a conformationally relatively rigid C-terminal heptapeptide and variable N-terminal fragments.     

Monte Carlo simulation of the conformational behaviour of a polypeptide chain near a charged surface

The conformational behaviour of a short polypeptide chain in the neighbourhood of a charged plane is simulated using a Monte Carlo method. In this approach, the plane is taken as a model of an interface separating a hydrophobic region from a hydrophilic one. It is shown that in the neighbourhood of the plane, folded molecular conformations are prevailing whilst stretched conformations are preferred far from the interface. When the plane is not charged, the molecule adjusts itself parallel to the interface. For a given position of the molecule with respect to the plane, when the charge density of the plane is increased, the molecule tends to turn perpendicular to the plane. The surface may either attract or repulse the molecule depending on the value of the charge density (the plane is always negatively charged).     

Local structure of cytochrome c from horse heart in solution. Conformational analysis using data of two-dimensional nuclear Overhauser effect spectroscopy]

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.     

Small molecule induction of MSH2-dependent cell death suggests a vital role of mismatch repair proteins in cell death

Avoidance of apoptosis is one of the hallmarks of cancer development and progression. Chemotherapeutic agents aim to initiate an apoptotic response, but often fail due to dysregulation. MSH proteins are capable of recognizing cisplatin damage in DNA and participate in the initiation of cell death. We have exploited this recognition and computationally simulated a MutS homolog (MSH) "death conformation". Screening and docking experiments based on this model determined that the MSH2-dependent cell-death pathway can be induced by a small molecule without DNA damage, reserpine. Reserpine was identified via virtual screening on structures obtained from molecular dynamics as a small molecule that selectively binds a protein "death" conformation. The virtual screening predicts that this small molecule binds in the absence of DNA. Cell biology confirmed that reserpine triggers the MSH2-dependent cell-death pathway. This result supports the hypothesis that the MSH2-dependent pathway is initiated by specific protein conformational changes triggered by binding to either DNA damage or small compound molecules. These findings have multiple implications for drug discovery and cell biology. Computational modeling may be used to identify and eventually design small molecules that selectively activate particular pathways through conformational control. Molecular dynamics simulations can be used to model the biologically relevant conformations and virtual screening can then be used to select for small molecules that bind specific conformations. The ability of a small molecule to induce the cell-death pathway suggests a broader role for MMR proteins in cellular events, such as cell-death pathways, than previously suspected.     

The multiple-minima problem in the conformational analysis of polypeptides. III. An Electrostatically Driven Monte Carlo Method: Tests on enkephalin

The three-dimensional conformation of Met-enkephalin, corresponding to the lowest minimum of the empirical potential energy function ECEPP/2 (empirical conformational energy program for peptides), has been determined using a new algorithm, viz. the Electrostatically Driven Monte Carlo Method. This methodology assumes that a polypeptide or protein molecule is driven toward the native structure by the combined action of electrostatic interactions and stochastic conformational changes associated with thermal movements. These features are included in the algorithm that produces a Monte Carlo search in the conformational hyperspace of the polypeptide, using electrostatic predictions and a random sampling technique to locate low-energy conformations. In addition, we have incorporated an alternative mechanism that allows the structure to escape from some conformational regions representing metastable local energy minima and even from regions of the conformational space with great stability. In 33 test calculations on Met-enkephalin, starting from arbitrary or completely random conformations, the structure corresponding to the global energy minimum was found inall the cases analyzed, with a relatively small search of the conformational space. Some of these starting conformations wereright orleft-handed -helices, characterized by good electrostatic interactions involving their backbone peptide dipoles; nevertheless, the procedure was able to convert such locally stable structures to the global-minimum conformation.On leave from the National University of San Luis, Faculty of Sciences and Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Instituto de Matemática Aplicada, San Luis, Ejército de los Andes 950, 5700 San Luis, Argentina.     

Conformation, and dissociation-recombination of chemically deglycosylated ovine lutropin

The conformations of intact and acid dissociated, as well as the relative rates of conformational repair of reassociating, ovine lutropin, and a chemically deglycosylated form of lutropin, have been studied by circular dichroism, zero-order, and difference absorption spectroscopy. Deglycosylation produces only small conformational changes in the intact or acid dissociated form of the molecule. While conformational repair of reassociating lutropin requires up to 72 h, and goes no further than 75-80%, deglycosylated lutropin reaches greater than 90% conformational repair in less than 4 h. The efficacy of second-order absorption spectra in characterizing conformations and conformational changes in these molecules is demonstrated.     

Theoretical study of prostaglandin E2 conformation

The theoretical conformational analysis of PGE2 has been performed using semiempirical approach and fixed values of the bond length and valence angles. A set of low-energy conformations was obtained for the alpha-chain and the whole PGE2 molecule. Unlike the PGE1 molecule, PGE2 alpha-chain prefers semifolded conformations. The hairpin structures with the parallel orientation of alpha- and omega-chains are the lowest-energy conformations of PGE2 conformations of PGE1 and PGE2. Sterically similar explain the similar biological activity of the two molecules.     

On the multiple-minima problem in the conformational analysis of polypeptides. IV. Application of the electrostatically driven Monte Carlo method to the 20-residue membrane-bound portion of melittin

The conformational space of the membrane-bound portion of melittin has been searched using the electrostatically driven Monte Carlo (EDMC) method with the ECEPP/2 (empirical conformational energy program for peptides) algorithm. The former methodology assumes that a polypeptide or protein molecule is driven toward the native structure by the combined action of electrostatic interactions and stochastic conformational changes associated with thermal movements. The algorithm produces a Monte Carlo search in the conformational hyperspace of the polypeptide using electrostatic predictions and a random sampling technique, combined with local minimization of the energy function, to locate low-energy conformations. As a result of 8 test calculations on the 20-residue membrane-bound portion of melittin, starting from six arbitrary and two completely random conformations, the method was able to locate a very low-energy region of the potential with a well-defined structure for the backbone. In all of the cases under study, the method found a cluster of similar low-energy conformations that agree well with the structure deduced from x-ray diffraction experiments and with one computed earlier by the build-up procedure.     

Disaccharide conformational flexibility. II. Molecular dynamics simulations of sucrose

Molecular dynamics simulations have been used to study the motions in vacuum of the disaccharide sucrose. Ensembles of trajectories were calculated for each of the five local minimum energy conformations identified in the adiabatic conformational energy mapping of this molecule. The model sucrose molecules were found to exhibit a variety of motions, although the global minimum energy conformation was found to be dynamically stable, and no transitions away from this structure were observed to occur spontaneously. In all but one of these vacuum trajectories, the intramolecular hydrogen bond between residues was maintained, in accord with recent nmr studies of this molecule in aqueous solution. Considerable flexibility of the furanoid ring was found in the trajectories. No "flips" to the opposite puckering for this ring were found in the simulations starting from the global minimum, although such a transition was observed for a trajectory initiated with one of the higher local minimum energy conformations. Overall, the observed structural fluctuations were consistent with the experimental picture of sucrose as a relatively rigid molecule.     

Hexapeptides That Inhibit Processing of Branched DNA Structures Induce a Dynamic Ensemble of Holliday Junction Conformations

Holliday junctions are critical intermediates in DNA recombination, repair, and restart of blocked replication. Hexapeptides have been identified that bind to junctions and inhibit various junction-processing enzymes, and these peptides confer anti-microbial and anti-tumor properties. Earlier studies suggested that inhibition results from stabilization of peptide-bound Holliday junctions in the square planar conformation. Here, we use single molecule fluorescence resonance energy transfer (smFRET) and two model junctions, which are AT- or GC-rich at the branch points, to show that binding of the peptide KWWCRW induces a dynamic ensemble of junction conformations that differs from both the square planar and stacked X conformations. The specific features of the conformational distributions differ for the two peptide-bound junctions, but both junctions display greatly decreased Mg²⁺ dependence and increased conformational fluctuations. The smFRET results, complemented by gel mobility shift and small angle x-ray scattering analyses, reveal structural effects of peptides and highlight the sensitivity of smFRET for analyzing complex mixtures of DNA structures. The peptide-induced conformational dynamics suggest multiple stacking arrangements of aromatic amino acids with the nucleobases at the junction core. This conformational heterogeneity may inhibit DNA processing by increasing the population of inactive junction conformations, thereby preventing the binding of processing enzymes and/or resulting in their premature dissociation.     

Quantitative determination of the conformation of ATP in aqueous solution using the lanthanide cations as nuclear-magnetic-resonance probes.

Chemical shift perturbations of the eight 1H resonances and of the three 31P resonances in the nuclear magnetic resonance spectra of ATP in 2H2O, pH 6.0, have been induced by specifically bound lanthanide cations Ln3+ (Ln = Pr, Nd, Eu, Yb). After separation of contact (through bond) perturbations the resultant through-space shifts, which are found to have axial symmetry, are used in an analysis of the conformation of the Ln3+ -ATP complex. A computer program was used to search for the conformations of the molecule which fit the nuclear magnetic resonance data. The "best" solutions obtained represent a small closely interrelated family of conformations. Effects of the cation Gd3+ on the longitudinal relaxation rates of five of the protons of ATP were also measured and used to confirm the conformational family. One of these conformations corresponds closely to one of the crystal structure forms, with an anti arrangement of the base-ribose unit and and a right-hand helical phosphate chain folded towards the adenine part of the molecule. The lanthanide ion binds predominantly to the beta and gamma phosphates and does not interact with the purine ring, these two centres being separated by at least one water molecule.     

Calculation of stable conformations of the molecule of a serum thymic factor

Stable conformations of "facteur thimique sérique" (FTS) were determined by semi-empirical conformational analysis. The interpretation of the calculation results in combination with data on biological activity of the conformationally restricted FTS analogues enabled the delineation of a possible "biologically active" conformation.     

Structure of valinomycin by molecular dynamics studies.

Valinomycin is an important ionophore which exhibits a high conformational flexibility. The study of various conformations adopted by this molecule together with the study of flexibility in a given conformation can throw light on the ion transport by the ionophore across the membrane. Molecular dynamics (MD) studies are ideal to characterize the flexibility in different parts of the molecule and can also give an idea of various conformations adopted by the molecule at a given temperature. Hence MD studies at 100K have been carried out on the minimized crystal structure of the molecule to scan the possible conformations in the neighbourhood of the well known 'bracelet' like structure of uncomplexed Valinomycin, Properties, like the flexibility, average values, r.m.s. fluctuations of the various intramolecular hydrogen bonds are discussed. Energy minimization has been carried out on selected MD simulated points to analyze the characteristics of the unique conformation adopted by this molecule at this temperature.     

Procedure for selecting starting conformations for energy minimization of nucleosides and nucleotides

The purpose of this study was to carry out a thorough search of the conformational space of various adenine-containing nucleotides, applying a previously published searching procedure, known as the representative method. This method, which reduces the number of starting conformations required to explore all the important regions of conformational space, appears to be successful in finding all (or nearly all) the putative low-energy conformations of each molecule.     

Examination of Matrix Metalloproteinase-1 in Solution: A PREFERENCE FOR THE PRE-COLLAGENOLYSIS STATE*

Catalysis of collagen degradation by matrix metalloproteinase 1 (MMP-1) has been proposed to critically rely on flexibility between the catalytic (CAT) and hemopexin-like (HPX) domains. A rigorous assessment of the most readily accessed conformations in solution is required to explain the onset of substrate recognition and collagenolysis. The present study utilized paramagnetic NMR spectroscopy and small angle x-ray scattering (SAXS) to calculate the maximum occurrence (MO) of MMP-1 conformations. The MMP-1 conformations with large MO values (up to 47%) are restricted into a relatively small conformational region. All conformations with high MO values differ largely from the closed MMP-1 structures obtained by x-ray crystallography. The MO of the latter is ∼20%, which represents the upper limit for the presence of this conformation in the ensemble sampled by the protein in solution. In all the high MO conformations, the CAT and HPX domains are not in tight contact, and the residues of the HPX domain reported to be responsible for the binding to the collagen triple-helix are solvent exposed. Thus, overall analysis of the highest MO conformations indicated that MMP-1 in solution was poised to interact with collagen and then could readily proceed along the steps of collagenolysis.     

Low-energy conformations of two lysine-containing tetrapeptides of collagen: implications for posttranslational lysine hydroxylation

The possible role of conformational constraints in the posttranslational hydroxylation of lysyl residues in collagen has been investigated by means of conformational energy computations for the N-acetyl-N′-methylamides of the four tetrapeptides Ala-Xxx-Gly-Ser and Gly-Xxx-Ala-Gly, where Xxx = Lys or Ala. When hydration is taken into account, all four peptides are shown to exist as a mixture of conformations, but there is a strong preference for type II bends in the conformational ensembles of two Ala-Xxx-Gly-Ser tetrapeptides and for type I bends in the conformational ensembles of the other two. The results agree with experimental evidence suggesting that a type II bend is an important conformation for Ac-Ala-Lys-Gly-Ser-NHCH₃, and they support an earlier suggestion that a β-bend may play a role in the posttranslational hydroxylation of Lys residues in position Y of the Gly-X-Y triplet in collagen.     

A shorter peptide model from staphylococcal nuclease for the folding-unfolding equilibrium of a beta-hairpin shows that unfolded state has significant contribution from compact conformational states

It is important to understand the conformational features of the unfolded state in equilibrium with folded state under physiological conditions. In this paper, we consider a short peptide model LMYKGQPM from staphylococcal nuclease to model the conformational equilibrium between a hairpin conformation and its unfolded state using molecular dynamics simulation under NVT conditions at 300K using GROMOS96 force field. The free energy landscape has overall funnel-like shape with hairpin conformations sampling the minima. The "unfolded" state has a higher free energy of approximately 12kJ/mol with respect to native hairpin minimum and occupies a plateau region. We find that the unfolded state has significant contributions from compact conformations. Many of these conformations have hairpin-like topology. Further, these compact conformational forms are stabilized by hydrophobic interactions. Conversion between native and non-native hairpins occurs via unfolded states. Frequent conversions between folded and unfolded hairpins are observed with single exponential kinetics. We compare our results with the emerging picture of unfolded state from both experimental and theoretical studies.