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At least 955 records · Page 53Linked to original sources
PubMed1996

Synthesis and Conformational Analysis of the Multidrug Resistance-Reversing Agent Hapalosin and Its Non-N-methyl Analog.

Hapalosin was initially synthesized by macrolactonization, and a second synthesis was achieved by cycloamidation. In both syntheses, three of the five stereocenters in hapalosin were established by two Brown allylboration reactions. The synthesis of the non-N-Me analog of hapalosin involved chelation-controlled reduction of a gamma-amino-beta-keto ester and cycloamidation. In CDCl(3) at 25 degrees C, synthetic hapalosin exists as a 2.3:1 mixture of conformers, while its non-N-Me analog exists only as a single conformer. (1)H,(1)H-NOESY and computation reveal that the configuration of the amide bond is responsible for the conformations of the two compounds. The major conformer of hapalosin is found to be an s-cis amide, the minor conformer an s-trans amide, and the non-N-Me analog an s-trans amide. Applying distance constraints to protons that exhibit NOESY correlations, computation shows that the major conformer of hapalosin and the non-N-Me analog have very different conformations. By contrast, the minor conformer of hapalosin and the non-N-Me analog have very similar conformations.

Journal Article↗
PubMed2001

The food mutagen 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline: a conformational analysis of its major DNA adduct and comparison with the 2-amino-3-methylimidazo[4,5-f]quinoline adduct.

The heterocyclic amine 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) is one of a group of heterocyclic amine carcinogens that exists in cooked meat and fish. It causes mutations in bacterial and mammalian assays and induces tumors in mammals. MeIQx is converted within cells to a reactive derivative which forms a major covalent adduct at carbon-8 of guanine in DNA. This adduct may alter the DNA conformation at critical stages of the replicative process, and cause mutations which initiate the carcinogenic process. Atomic resolution structures of the MeIQx-damaged DNA are not yet available experimentally. We have carried out an extensive molecular mechanics/energy minimization search to locate feasible structures for the major MeIQx adduct in DNA, using the sequence d(5'-C1-G2-C3-G4[IQ]-C5-G6-C7-3').d(5'-G8-C9-G10-C11-G12-C13-G14-3') with MeIQx modification at G4. We have created 1152 starting conformations which uniformly sampled each of the three flexible torsion angles that govern the MeIQx-DNA orientation at 15 degrees intervals, and minimized their energy. A mixture of conformations was generated, which were separated into families according to the position of the ring system of the carcinogenic amine: major groove, minor groove, and base-displaced-intercalated. While a generally similar mixture had been generated previously for the related carcinogen 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) [Wu, X., et al. (1999) Chem. Res. Toxicol. 12, 895-905], differences were found which could be rationalized in terms of the additional methyl group in the MeIQx.

Animals↗
PubMed2005

Magic-angle spinning solid-state NMR spectroscopy of the beta1 immunoglobulin binding domain of protein G (GB1): 15N and 13C chemical shift assignments and conformational analysis.

Magic-angle spinning solid-state NMR (SSNMR) studies of the beta1 immunoglobulin binding domain of protein G (GB1) are presented. Chemical shift correlation spectra at 11.7 T (500 MHz 1H frequency) were employed to identify signals specific to each amino acid residue type and to establish backbone connectivities. High sensitivity and resolution facilitated the detection and assignment of every 15N and 13C site, including the N-terminal (M1) 15NH3, the C-terminal (E56) 13C', and side-chain resonances from residues exhibiting fast-limit conformational exchange near room temperature. The assigned spectra lend novel insight into the structure and dynamics of microcrystalline GB1. Secondary isotropic chemical shifts report on conformation, enabling a detailed comparison of the microcrystalline state with the conformation of single crystals and the protein in solution; the consistency of backbone conformation in these three preparations is the best among proteins studied so far. Signal intensities and line widths vary as a function of amino acid position and temperature. High-resolution spectra are observed near room temperature (280 K) and at <180 K, whereas resolution and sensitivity greatly degrade substantially near 210 K; the magnitude of this effect is greatest among the side chains of residues at the intermolecular interface of the microcrystal lattice, which we attribute to intermediate-rate translational diffusion of solvent molecules near the glass transition. These features of GB1 will enable its use as an excellent model protein not only for SSNMR methods development but also for fundamental studies of protein thermodynamics in the solid state.

Amino Acid Sequence↗
PubMed1987

Conformational analysis of a cyclic thymopoietin-analogue by 1H n.m.r. spectroscopy and restrained molecular dynamics simulations.

The internuclear distances of the cyclic thymopoietin derivative c[D-Val-Tyr-Arg-Lys-Glu] have been determined using two-dimensional nuclear Overhauser n.m.r. spectroscopy. These distances are used as constraints for a restrained Molecular Dynamics (MD) simulation. The two starting structures used for the calculations consist of a beta and gamma turn for model 1 and two gamma turns for model 2. The rms difference in atomic positions of the two conformations is 0.242 nm. They converge during the restrained MD simulation to the same final structure. The positional rms difference of the time averaged (5-14 ps) conformations is 0.011 nm. The hydrogen bond pattern is similar to that of model 1, but in addition we find three more gamma turns. The vicinal NH-C alpha H couplings agree well with those calculated from the time averaged structures.

Magnetic Resonance Spectroscopy↗
PubMed2006

Conformational analysis of single DNA molecules undergoing entropically induced motion in nanochannels.

We have used the interface between a nanochannel and a microchannel as a tool for applying controlled forces on a DNA molecule. A molecule, with a radius of gyration larger than the nanochannel width, that straddles such an interface is subject to an essentially constant entropic force, which can be balanced against other forces such as the electrophoretic force from an applied electric field. By controlling the applied field we can position the molecule as desired and observe the conformation of the molecule as it stretches, relaxes, and recoils from the nanochannel. We quantify and present models for the molecular motion in response to the entropic, electrophoretic, and frictional forces acting on it. By determining the magnitude of the drag coefficients for DNA molecules in the nanostructure, we are able to estimate the confinement-induced recoil force. Finally, we demonstrate that we can use a controlled applied field and the entropic interfacial forces to unfold molecules, which can then be manipulated and positioned in their simple extended morphology.

DNA↗
PubMed1998

Novel 3'-spiro nucleoside analogues of TSAO-T. Part II. A comparative study based on NMR conformational analysis in solution and theoretical calculations.

The structures of two novel 3'-spiro nucleosides analogues of the potent human immunodeficiency virus type 1 (HIV-1) reverse trancriptase (RT) inhibitor TSAO-m3T, in solution, as derived from NMR spectroscopy are described. In these TSAO analogues the spiro amino oxathioledioxide moiety has been replaced by spiro amino oxazolone or spiro amino oxathiazoledioxide moieties. A comparative study based on theoretical calculations of the hydrophobicity, the solvation free energies and molecular electrostatic potentials (MEP) of the three compounds is also described. No significant conformational differences were detected in solution between TSAO-m3T and its analogues that might account for the differences observed in their inhibitory activity against HIV-1 RT. The calculated hydrophobicity (log P) values, dipole moments and the electrostatic contributions to the solvation free energies of the three spiro ring systems were also similar. However, the differences found in the calculated MEPs of the spiro systems between TSAO-m3T and its analogues suggest that the different electrostatic surroundings of the 4"-amino group of the spiro moiety in the analogues may be responsible for a detrimental electrostatic interaction of the spiro rings with the Glu-B138 of RT.

Anti-HIV Agents↗
PubMed1992

Conformational analysis of Met-enkephalin in both aqueous solution and in the presence of sodium dodecyl sulfate micelles using multidimensional NMR and molecular modeling.

Proton and 13C chemical shift assignments are reported for the neuropeptide Met-enkephalin (ME) in both aqueous solution and in the presence of 50 mM sodium dodecyl sulfate (SDS). Rotating frame nuclear Overhauser enhancement spectroscopy was used to qualitatively describe interproton distances. These distances were then used as restraints in the distance geometry based molecular modeling program Dspace, developed by Hare Research to generate sets of conformations of ME. The resulting aqueous solution conformations of ME were determined to exhibit characteristic of an extended random-coil polypeptide with no distinguishable secondary structure. The resulting set of solution conformations of ME in the presence of 50 mM SDS exhibited characteristics of an amphiphilic type IV beta turn that are stabilized by hydrophobic aromatic-aromatic interactions between the side chains of Tyr1 and Phe4.

Amino Acid Sequence↗
PubMed1983

Conformational analysis of oligoarabinonucleotides. An NMR and CD study.

A 500 and 300 MHz proton NMR study of the series of oligoarabinonucleotides 5'aAMP, 3'aAMP, aA-aA, (aA-)2aA and (aA-)3aA is presented. In addition, circular dichroism is used to study the stacking behaviour of aA-aA. The complete 1H-NMR spectral assignment of the compounds (except the tetramer) is given. Proton-proton and proton-phosphorus coupling constants, obtained by computer simulation of the high-field region of the spectra, yield information on the conformation of the arabinose rings (N- or S-type) and on the intramolecular stacking properties of the dimer and the trimer. The monomers 5'aAMP and 3'aAMP exhibit a preference for N- and S-type sugar conformation, respectively. It is shown that the dimer aA-aA at low temperature prefers a mixed stacked state of the type aA(S)-aA(N). In the trimer the aA(2)-aA(3) fragment exhibits a conformation similar to that found in the dimer, whereas the aA(1) residue prefers to adopt S-type sugar and has some tendency to stack upon residue aA(2).

Arabinonucleotides↗
PubMed1993

Conformational analysis and proteolytic processing of synthetic pre-pro-GnRH/GAP protein.

Homogeneous pre-pro-GnRH/GAP protein was recently synthesized in 100 mg quantities by solid-phase methods and surprisingly, the synthetic pre-pro-protein, which normally does not escape the endoplasmic reticulum, was found to inhibit the release of prolactin from cultured pituitary cells. This is the first demonstration of significant biological activity associated with a precursor protein and provides the rationale for its further study. We now report the results of our initial examination of the conformational properties of pre-pro-GnRH/GAP protein as a prelude to solving its solution phase conformation by homonuclear 1H-NMR protocols. Thermal and pH titration fluorescence and circular dichroism spectroscopies reveal that the protein is resistant to thermal-induced conformational changes but is particularly sensitive to pH-induced conformational changes; while Asp/Glu and Arg residues may contribute to structural stability, His and Lys residues predominate. Pre-pro-GnRH/GAP is about 30% helix in the range of 2-40 degrees C; however, even at 90 degrees C, the peptide retains nearly 50% of its helix character. There is no evidence for a cooperative transition; for this reason, differential scanning calorimetry failed to yield a defined transition thermogram. Pre-pro-GnRH/GAP apparently does not pass through a transition state as a function of temperature but appears to flex and retain a high percentage of helix structure, resulting in subtle changes in secondary structure. There is no discernible isodichroic point. On either side of the neutral pH range, however, there are dramatic changes in structure that result in nonreversible denaturation of the protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed2002

Intramolecular radical cyclization-ring-closing metathesis approach to fused polycyclic ethers. Convergent synthesis and conformational analysis of the (E)FGH ring system of ciguatoxin.

A convergent synthetic route to the (E)FGH ring system 4 of ciguatoxins, the causative toxins for ciguatera fish poisoning, has been developed. The synthesis features convergent coupling to form dioxane acetal, regioselective acetal cleavage by diethylaluminum phenylthiolate or diisobutylaluminum phenylselenolate followed by intramolecular radical cyclization to construct the oxepane ring G, and a ring-closing metathesis reaction to form the hexahydrooxonine ring F. The hexahydrooxonine ring F of tetracyclic model system 4 existed as a 5:1 equilibrium mixture of two conformers (UP and DOWN conformers), with the UP one predominating. This is the first illustration that reproduces the preference for the UP conformer over the DOWN one, which preference was observed for natural ciguatoxins.

Acetals↗
PubMed2002

Spirolactams as conformationally restricted pseudopeptides: synthesis and conformational analysis.

The synthesis of 1-(tert-butoxycarbonyl)-7-[1-(tert-butoxycarbonyl)-3-methylbutyl]-6-oxo-1,7-diazaspiro[4.5]decanes (S,S)-1a and (S,R)-1b is described. Derivatives 17a,b and 19a are prepared for use in peptide synthesis as constrained surrogates of the Pro-Leu and Gly-Leu dipeptides. The Ac-[Gly-Leu]-Met-NH(2) derivatives (S,S,S)-2a and (S,R,S)-2b, with the tripeptidic C-terminal region present in tachykinins, are also synthesized. Conformational analyses of these tripetide analogues by NMR experiments and molecular modeling calculations show that both (S,S,S)-2a and (S,R,S)-2b epimers are gamma-turn/distorted type II beta-turn mimetics.

Hydrogen Bonding↗
PubMed1992

Neuropeptide Y. Optimized solid-phase synthesis and conformational analysis in trifluoroethanol.

The 36-amino-acid neuropeptide Y (human), which is one of the most potent vasoconstrictors and which exhibits a number of other biological functions, has been synthesized using automated peptide synthesis. The optimized method, using 9-fluorenylmethoxycarbonyl protecting and single-step coupling, yielded the crude product in 90% purity allowing for single-step reversed-phase HPLC purification to greater than 98% purity and a high overall yield (50%). The hormone was characterized by several chromatographic methods, ion-spray mass spectroscopy and Edman degradation. The conformation of human neuropeptide Y was examined by CD, NMR and computer simulations. The CD measurements in trifluoroethanol/water (9:1) show a large percentage of alpha-helix. Variation of concentration, from 0.5 microM increasing up to the 1 mM used for NMR measurements, indicates no evidence for aggregation. In the same solvent system, the NMR line widths were very broad and therefore the resonance assignment was achieved with the exclusive use of two-dimensional NOE spectra. The 248 clearly distinguishable NOEs from the NMR study were used in distance geometry calculations and the resulting structures were refined with restrained molecular dynamics. The results indicate an alpha-helix extending from Arg19 to Gln34. For the N-terminal half of the molecule no regular structure was observed.

Amino Acid Sequence↗
PubMed2004

Conformational analysis of hepatitis B surface antigen fusions in an Agrobacterium-mediated transient expression system.

Vaccine antigens have been successfully produced in transgenic plants for oral immunization. Recently, a fusion strategy has been adopted to produce multicomponent vaccines and to target antigens to mucosal sites for enhanced oral immunogenicity. However, antigen fusions may not be folded correctly due to steric hindrance and may thus lose their potency. Here, we describe an Agrobacterium-mediated transient assay that provides enough antigen-expressing material at 2 days post-transfection to evaluate antigen conformation. Using the hepatitis B surface antigen (HBsAg) as a model antigen and the green fluorescent protein (GFP) as a model fusion partner, we showed that transiently expressed HBsAg and an HBsAg fusion with GFP at the N-terminus (GFP:HBsAg), but not the HBsAg fusion with GFP at the C-terminus (HBsAg:GFP), formed the 'a' determinant and virus-like particles (VLPs), similar to yeast-derived vaccine HBsAg. Thus, it is feasible to modify the HBsAg with an N-terminal fusion of up to 239 amino acids without altering its major antigenic properties. Our results also demonstrate that the Agrobacterium-mediated transient expression system can be used to evaluate the conformation of plant-based vaccines or other pharmaceutical proteins in a high-throughput manner.

Journal Article↗
PubMed1994

The conformational analysis of a synthetic S4 peptide corresponding to a voltage-gated potassium ion channel protein.

The S4 region of the Drosophila Shaker voltage-gated K+ channel has been proposed to function as a voltage-sensor. We have synthesised a peptide corresponding to this S4 region. Structural studies on the S4 peptide were conducted using Fourier transform infrared (FTIR) spectroscopy. Spectra were obtained for the peptide dissolved in aqueous solution, in trifluoroethanol solvent and also after reconstitution into lipid bilayers and micelles. The peptide in trifluoroethanol adopts an alpha-helical conformation which is in good agreement with the results of a recent 2D NMR study on the structure of a S4 peptide corresponding to the rat brain sodium channel [(1989) FEBS Lett. 257, 113-117]. A predominantly alpha-helical structure is also observed when the S4 peptide is present in aqueous lysophosphatidylcholine micelles, in dimyristoyl phosphatidylcholine and dimyristoyl phosphatidylglycerol lipid bilayers. In contrast to this, the S4 peptide in aqueous solution is in a random coil conformation. The coil-to-helix transition observed for the S4 peptide upon its transfer from aqueous solution to lipid membrane indicates that it has a high degree of conformational flexibility and can undergo large changes in its structure in response to its environment. This may have important implications for its role in the voltage activation process during which the S4 peptide has been postulated to, at least partially, move from a lipid bilayer to an aqueous extracellular media [(1992) Biophys J. 62, 238-250]. The results of our study lend support to such a model.

Amino Acid Sequence↗
PubMed1989

Structure and conformational analysis of the opioid antagonist (-)-(1R,5R,9R)-5,9-diethyl-2-(3-furylmethyl)-2'-hydroxy-6,7-benzomorpha n (Mr2266).

C21H27NO2, Mr = 325.449, monoclinic, P2(1), a = 16.3916 (7), b = 12.7460 (5), c = 8.9806 (5) A, beta = 107.191 (4) degrees, V = 1792.5 (2) A3, Z = 4, Dm = 1.22 (2), D chi = 1.206 Mg m-3, lambda(Cu K alpha) = 1.54178 A, mu(Cu K alpha) = 0.566 mm-1, F(000) = 704, T = 291 K, final R = 0.048 for 4225 observed reflections. The two molecules present in the asymmetric unit adopt a different conformation with respect to the N-side chain. Starting from the asymmetric carbon and proceeding along the allyl moiety the conformations are antiperiplanar/(-)-anticlinal for molecule 1 and antiperiplanar/(+)-synclinal for molecule 2. The furyl rings engage in aromatic-aromatic interactions which are compared with results from a theoretical study from the literature. Finally, the 3-furyl geometry is evaluated through a Cambridge Structural Database search and CNDO/2 calculations.

Benzomorphans↗
PubMed1977

A conformational analysis for leucine-enkephalin using activity and binding data of synthetic analogues.

1. Leucine-enkephalin and some analogues were assayed for activity in vitro on the mouse vas deferens and for binding to opiate receptors from rat brain. 2. The experimental data were analysed in terms of the stringency for glycine, a D-amino acid or an L-amino acid at each position in the peptide. 3. The observed configurational specificity was compared with the stringency that would be predicted to occur if enkephalin adopted certain hydrogen-bonded conformations at the receptor. 4. A small subset of the conformations examined was found to be compatible with the experimental data.

Animals↗
PubMed1993

Conformational analysis of human dihydrofolate reductase inhibitor complexes: crystal structure determination of wild type and F31 mutant binary and ternary inhibitor complexes.

These structural studies reveal unusual intermolecular interactions for the binding of inhibitors and cofactor in ternary complexes with both wild type and F31 mutant recombinant human DHFR and show that these inhibitors have flexibility in occupying the active site. These studies also possibly indicate the first structural data for a ternary complex with a folate inhibitor and a polyglutamate side chain. However, further refinement of this data is necessary before this can be confirmed. In contrast to the ternary complexes of folate and MTX, the lipophilic antifolate PTX binds with its methoxybenzoyl ring oriented toward the cofactor nicotinamide ring, while that of TMQ it is bound closer to the Phe-31 position. Furthermore, the nicotinamide ring makes a close contact to the N10 amine of TMQ, significantly different from its binding site interactions in MTX complexes. These data also reveal that the conserved contacts between the cofactor carboxyamide with the enzyme backbone residues Ala-9 and Ile-16 are dictated by the enzyme and that changes in the orientation of the structural elements requires only subtle changes in the secondary structural units in which they are contained. Therefore, only by careful analysis of a series of enzyme complexes can the mechanisms of binding action be delineated.

Binding Sites↗