Synthesis and conformational analysis of novel N(OCH3)-linked disaccharide analogues

N(OMe)-linked disaccharide analogues, isosteric to the corresponding natural disaccharides, have been synthesized by chemoselective assembly of unprotected natural monosaccharides with methyl 6-deoxy-6-methoxyamino-alpha-D-glucopyranoside in an aqueous environment. The coupling reactions were found to be chemo- and stereoselective affording beta-(1-->6) disaccharide mimics when using Glc and GlcNAc; in the case of Gal, the beta-anomer was prevalent (beta:alpha=7:1). An iterative method for the synthesis of linear N(OMe) oligosaccharide analogues was demonstrated, based on the use of an unprotected monosaccharide building block in which an oxime functionality at C-6 is converted during the synthesis into the corresponding methoxyamino group. The conformational analysis of these compounds was carried out by using NMR spectroscopy, ab initio, molecular mechanics, and molecular dynamics methods. Optimized geometries and energies of fourteen conformers for each compound have been calculated at the B3LYP/6-31G* level. Predicted conformational equilibria were compared with the results based on NMR experiments and good agreement was found. It appears that N(OMe)-linked disaccharide analogues exhibit a slightly different conformational behavior to their parent natural disaccharides.     

The N‐Terminal Nonapeptide of Cephaibols A and C: A Naturally Occurring Example of Mismatched Helical Screw‐Sense Control

The N‐terminal nonapeptide domain of the fungal nonribosomal peptide antibiotics cephaibol A and cephaibol C (AcPheAib₄LeuIvaGly‐ Aib) is reported to adopt a right‐handed helical conformation in the crystalline state. However, this conformation is at odds with the left‐handed helicity observed in solution in related synthetic oligomers capped with Ac‐L ‐PheAib₄ fragments. We report the synthesis of four diastereoisomers of the cephaibol N‐terminal nonapeptide, and show by NMR and CD spectroscopy that the peptide containing the chiral amino acids Phe and Leu in the naturally occurring relative configuration exists in solution as an interconverting mixture of helical screw‐sense conformers. In contrast, the nonapeptide containing the unnatural relative configuration at Phe and Leu adopts a single, stable helical screw‐sense, which is left handed when the N‐terminal Phe residue is L and right‐handed when the N‐terminal Phe residue is D .     

1H NMR spectroscopic studies of the conformational isomers of pyrrolidinofullerenes

Mixed bis-adduct derivatives of C60 containing a pyrrolidine and a malonate methano group were synthesized. Three regioisomers, the e', the trans-2, and the trans-3, were isolated and characterized. In-depth NMR studies of these methano-pyrrolidinofullerenes showed that the nitrogen inversion on the pyrrolidine moiety is not a fast event in the 1H NMR time scale as previously regarded. Solvent effects, variable temperature experiments, and protonation of the pyrrolidine nitrogen are addressed.     

Cyclic beta-tetra- and pentapeptides: synthesis through on-resin cyclization and conformational studies by X-ray, NMR and CD spectroscopy and theoretical calculations

The solution-phase synthesis of the simplest cyclic beta-tetrapeptide, cyclo(beta-Ala)4 (4), as well as the solid-phase syntheses through side chain anchoring and on-resin cyclization of the cyclic beta3-tetrapeptide cyclo(-beta3hPhe-beta3hLeu-beta3hLys-beta3hGln-) (14) and the first cyclic beta3-pentapeptide cyclo(-beta3hVal-beta3hPhe-beta3hLeu-beta3hLys-beta3hLys-) (19) are reported. Extensive computational as well as spectroscopic studies, including X-ray and NMR spectroscopy, were undertaken to determine the preferred conformations of these unnatural oligomers in solution and in the solid state. cyclo(beta-Ala)4 (4) with no chiral side chains is shown to exist as a mixture of rapidly interchanging conformers in solution, whereas inclusion of chiral side chains in the cyclo-beta3-tetrapeptide causes stabilization of one dominating conformer. The cyclic beta3-pentapeptide on the other hand shows larger conformational freedom. The X-ray structure of achiral cyclo(beta-Ala)4 (4) displays a Ci-symmetrical 16-membered ring with adjacent C=O and N-H atoms pointing pair wise up and down with respect to the ring plane. CD spectroscopic examinations of all cyclic beta-peptides were undertaken and revealed results valuable as starting point for further structural investigations of these entities.     

Chemical modulation of peptoids: synthesis and conformational studies on partially constrained derivatives

The high conformational flexibility of peptoids can generate problems in biomolecular selectivity as a result of undesired off-target interactions. This drawback can be counterbalanced by restricting the original flexibility to a certain extent, thus leading to new peptidomimetics. By starting from the structure of an active peptoid as an apoptosis inhibitor, we designed two families of peptidomimetics that bear either 7-substituted perhydro-1,4-diazepine-2,5-dione 2 or 3-substituted 1,4-piperazine-2,5-dione 3 moieties. We report an efficient, solid-phase-based synthesis for both peptidomimetic families 2 and 3 from a common intermediate. An NMR spectroscopic study of 2a,b and 3a,b showed two species in solution in different solvents that interconvert slowly on the NMR timescale. The cis/trans isomerization around the exocyclic tertiary amide bond is responsible for this conformational behavior. The cis isomers are more favored in nonpolar environments, and this preference is higher for the six-membered-ring derivative 3a,b. We propose that the hydrogen-bonding pattern could play an important role in the cis/trans equilibrium process. These hydrogen bonds were characterized in solution, in the solid state (i.e., by using X-ray studies), and by molecular modeling of simplified systems. A comparative study of a model peptoid 10 containing the isolated tertiary amide bond under study outlined the importance of the heterocyclic moiety for the prevalence of the cis configuration in 2a and 3a. The kinetics of the cis/trans interconversion in 2a, 3a, and 10 was also studied by variable-temperature NMR spectroscopic analysis. The full line-shape analysis of the NMR spectra of 10 revealed negligible entropic contribution to the energetic barrier in this conformational process. A theoretical analysis of 10 supported the results observed by NMR spectroscopic analysis. Overall, these results are relevant for the study of the peptidomimetic/biological-target interactions.     

Non-natural amino acids as modulating agents of the conformational space of model glycopeptides

The synthesis and conformational analysis in aqueous solution of different alpha-methyl-alpha-amino acid diamides, derived from serine, threonine, beta-hydroxycyclobutane-alpha-amino acids, and their corresponding model beta-O-glucopeptides, are reported. The study reveals that the presence of an alpha-methyl group forces the model peptides to adopt helix-like conformations. These folded conformations are especially significant for cyclobutane derivatives. Interestingly, this feature was also observed in the corresponding model glucopeptides, thus indicating that the alpha-methyl group and not the beta-O-glucosylation process largely determines the conformational preference of the backbone in these structures. On the other hand, atypical conformations of the glycosidic linkage were experimentally determined. Therefore, when a methyl group was located at the Cbeta atom with an R configuration, the glycosidic linkage was rather rigid. Nevertheless, when the S configuration was displayed, a significant degree of flexibility was observed for the glycosidic linkage, thus showing both alternate and eclipsed conformations of the psi(s) dihedral angle. In addition, some derivatives exhibited an unusual value for the phi(s) angle, which was far from a value of -60 degrees expected for a conventional beta-O-glycosidic linkage. In this sense, the different conformations exhibited by these molecules could be a useful tool in obtaining systems with conformational preferences "à la carte".     

Synthesis and conformational analysis of glycomimetic analogs of thiochitobiose

The synthesis of six analogs of N,N′-diacetylchitobiose is reported, including a novel transglycosylation reaction for the preparation of S-aryl thioglycosides. The conformations of the compounds were studied by a combination of NMR spectroscopy and molecular modeling, using force field calculations. In the case of the S-aryl thioglycosides with exclusively S-glycosidic linkages, dihedral angles of the disaccharidic S-glycosidic bonds, Φ′ and Ψ′ and of the S-arylglycoside bonds, Φ and Ψ, were found to be similar, whereas they were different in mixed glycosides and in a thiazoline derivative. An adequate correlation between the calculated H,H-distances of the local minima and the measured NOE contacts was achieved by applying population-weighted averages over participating conformers based on weighted relative energies.     

Solution- and bound-state conformational study of N,N',N"-triacetyl chitotriose and other analogous potential inhibitors of hevamine: application of trNOESY and STD NMR spectroscopy

The solution-state conformations of N,N',N"-triacetyl chitotriose (1) and other potential chitinase inhibitors 2-4 were studied using a combination of NMR spectroscopy (NOESY) and molecular mechanics calculations. Determination solely of the global energy minimum conformation was found to be insufficient for an agreement with the NMR results. An appropriate consistency between the NMR experimental data and theoretical calculations was only reached by assessing the structures as population-weighted average conformers based on Boltzmann distributions derived from the calculated relative energies. Analogies, but also particular differences, between the synthetic compounds 2-4 and the naturally-occurring N,N',N"-triacetyl chitotriose were found. Furthermore, the conformation of compounds 1 and 2 when bound to hevamine was also studied using transferred NOESY experiments and the binding process was found to impart a level of conformational restriction on the ligands. The preferred conformation as determined for 1 in the bound state to hevamine belonged to one of the conformational families found for the compound when free in solution, although full characterisation of the bound-state conformations was impeded due to severe signal overlap. Saturation transfer difference NMR experiments were also employed to analyse the binding epitopes of the bound compounds. We thus determined that it is mainly the acetyl amido groups of the trisaccharide and the heterocyclic moiety which are in close contact with hevamine.     

Electronic and steric substituent influences on the conformational equilibria of cyclohexyl esters: the anomeric effect is not anomalous!

The cyclohexyl esters of a series of carboxylic acids, RCO₂H, spanning a range of electronegativities and quotients of steric hindrance for the R substituent (R=Me, Et, iPr, tBu, CF₃, CH₂Cl, CHCl₂, CCl₃, CH₂Br, CHBr₂, and CBr₃) were prepared. Their conformational equilibria in CD₂Cl₂ were examined by low‐temperature ¹H NMR spectroscopy to study the axial or equatorial orientation of the ester functionality with respect to the adopted chair conformation of the cyclohexane ring. The ab initio and DFT geometry‐optimized structures and relative free energies of the axial and equatorial conformers were also calculated at the HF/6‐311G**, MP2/6‐311G**, and B3LYP/6‐31G** levels of theory, both in the gas phase and in solution. In the latter case, a self‐consistent isodensity polarized continuum model was employed. Only by including electron correlation in the modeling calculations for the solvated molecules was it possible to obtain a reasonable correlation between ΔG°_calcd and ΔG°_exp. Both the structures and the free energy differences of the axial and equatorial conformers were evaluated with respect to the factors normally influencing conformational preference, namely, 1,3‐diaxial steric interactions in the axial conformer and hyperconjugation. It was assessed that hyperconjugative interactions, σ_{C? C}/σ_{C? H} and σ*_{C? O}, together with a steric effect—the destabilization of the equatorial conformer with increasing bulk of the R group—were the determinant factors for the position of the conformational equilibria. Thus, because hyperconjugation is held responsible as the mitigating factor for the anomeric effect in 2‐substituted, six‐membered saturated heterocyclic rings, and since it is also similarly responsible, at least partly, in these monosubstituted cyclohexanes for a preferential shift towards the axial conformer, the question is therefore raised: can the anomeric effect really be construed as anomalous?     

Simultaneous definition of high resolution protein structure and backbone conformational dynamics using NMR residual dipolar couplings.

Despite the importance of molecular dynamics for biological activity, most approaches to protein structure determination, whether based on crystallographic or solution studies, propose three-dimensional atomic representations of a single configuration that take no account of conformational fluctuation. Non-averaged anisotropic NMR interactions, such as residual dipolar couplings, that become measurable under conditions of weak alignment, provide sensitive probes of both molecular structure and dynamics. Residual dipolar couplings are becoming increasingly powerful for the study of proteins in solution. In this minireview we present their use for the simultaneous determination of protein structure and dynamics.     

Unexpected conformational properties of 1-trifluoromethyl-1-silacyclohexane, C5H10SiHCF3: gas electron diffraction, low-temperature NMR spectropic studies, and quantum chemical calculations

The molecular structure of axial and equatorial conformers of 1-trifluoromethyl-1-silacyclohexane, (C5H10SiHCF3), as well as the thermodynamic equilibrium between these species was investigated by means of gas electron diffraction (GED), dynamic nuclear magnetic resonance (DNMR) spectroscopy, and quantum chemical calculations (B3LYP, MP2, and CBS-QB3). According to GED, the compound exists as a mixture of two Cs symmetry conformers possessing the chair conformation of the six-membered ring and differing in the axial or equatorial position of the CF3 group (axial=58(12) mol%/equatorial=42(12) mol%) at T=293 K. This result is in a good agreement with the theoretical prediction. This is, however, in sharp contrast to the conformational properties of the cyclohexane analogue. The main structural feature for both conformers is the unusually long exocyclic bond length Si--C 1.934(10) A. A low-temperature 19F NMR experiment results in an axial/equatorial ratio of 17(2) mol%:83(2) mol% at 113 K and a DeltaG (not equal) of 5.5(2) kcal mol-1. CBS-QB3 calculations in the gas-phase and solvation effect calculations using the PCM(B3LYP/6-311G*) and IPCM(B3LYP/6-311G*) models were applied to estimate the axial/equatorial ratio in the 100-300 K temperature range, which showed excellent agreement with the experimental results. The minimum energy pathways for the chair-to-chair inversion of trifluoromethylsilacyclohexane and methylsilacyclohexane were also calculated using the STQN(Path) method.     

Synthesis and conformational analysis of 18-membered Aib-containing cyclohexapeptides

The synthesis and conformational analysis of two Aib-containing cyclic hexapeptides, cyclo(Gly-Aib-Leu-Aib-Phe-Aib) 1 and cyclo(Leu-Aib-Phe-Gly-Aib-Aib) 2, is described. The linear precursors of 1 and 2 were prepared using solution phase techniques, and the cyclization efficiency of three different coupling reagents (HATU, PyAOP, DEPC) was examined. The success of the cyclization was found to be reagent dependent. Solid-state conformational analysis of 1 and 2 was performed by X-ray crystallography and has revealed some unusual features as all three Aib residues of 1 assume nonhelical conformations. Furthermore, the residue Aib⁴ adopts an extended conformation (?=−175.9(3)°, ψ=+178.6(2)°), which is, to the best of our knowledge, the first observation of an Aib residue adopting an extended conformation in a cyclopeptide. The structure of 1 is also a rare example in which an Aib residue occupies the (i+1) position of a type II′ β-turn, stabilized by a bifurcated hydrogen bond. The cyclic peptide 2 adopts a more regular conformation in the solid state, consisting of two fused β-turns of type I/I′, stabilized by a pair of intramolecular hydrogen bonds. In addition, the conformational study of the cyclic peptide 1 in DMSO-d₆ by NMR spectroscopy and molecular dynamics simulations revealed a structure, which is very similar to its structure in the crystalline state.     

Synthesis and conformational behaviour of ditosyldiaza[2.2]orthometacyclophanes

Summary The N,N-ditosyl-diaza[2.2]orthometacyclophanes5a, b were synthesized from N,N-ditosyl-metaphenylenediamine by reaction with (Z)-1,4-dichlorobutene and 1,2-bis-bromomethylbenzene, respectively. Low temperature NMR studies showed that the compound5b exists as a 1:1 mixture of chair and boat form of the strained (E,Z)-diazanonadiene ring. At room temperature all corresponding resonances are averaged on the NMR time scale (including all four ethylene bridge protons). Going to lower temperatures, in a first step the methylene bridge inversion is frozen (giving twoexo H and twoendo H, G ca. 52kJmol^–1). In a second step the chair and boat form can be observed separately (G ca. 43.5 kJ mol^–1 for the chair/boat flip). The assignments were confirmed by 2D NMR experiments.Dedicated to Prof. Dr. E. L. Eliel on the occasion of his 70th anniversary     

Intrinsic information content of NMR dipolar couplings: a conformational investigation of 1,3-butadiene in a nematic phase.

The conformational equilibrium of 1,3-butadiene in a condensed fluid phase is investigated by liquid-crystal NMR spectroscopy. The full set of D(HH) and D(CH) dipolar couplings is determined from the analysis of the (1)H spectra of the three 1,3-butadiene most-abundant isotopomers (i.e. the all (12)C and the two single-labeled (13)C isotopomers) for a total of 21 independent dipolar couplings. A very good starting set of spectral parameters for the analysis of the (1)H spectrum is determined in a semiautomated way by the analysis of the (N-1) (specifically, N=6, the number of 1/2 spin nuclei in the spin system) quantum refocused (5QR), and not (5Q), spectra. As an alternative approach, a Monte Carlo (MC) numerical simulation, capable of predicting the solute ordering, is tested to simulate the 5QR spectrum. The set of D(ij) couplings is very good, proving that the MC method can represent a novel, valid alternative to the existing spectral simplification procedures. The experimentally determined dipolar-coupling data set is fully compatible with the 1,3-butadiene conformational distribution reported in the literature for isolated molecules, indicating the presence of about 99 % of s-trans conformer. With regards to the remaining 1 %, in spite of the direct and very strong dependence of the observables on the molecular structure, it was not possible to discriminate between the planar s-cis and s-gauche forms, both of which produce a very good fit of the dipolar couplings. Vibrational corrections, up to the anharmonic term, were applied; the calculated geometrical parameters are in good- although not exact-agreement with those reported in the literature from experimental and theoretical investigations. This result can be considered as supporting the methodology used for obtaining the structure and conformational distribution of a flexible molecule in a liquid phase.