Vicinal 1H-1H Coupling Constants Research Articles

Dactylides A-C, three new bioactive 22-membered macrolides produced by Dactylosporangium aurantiacum.

Three new 22-membered polyol macrolides, dactylides A-C (1-3), were isolated from Dactylosporangium aurantiacum ATCC 23491 employing repeated chromatographic separations, and their structures were established based on detailed analysis of NMR and MS data. The relative configurations at the stereocenters were established via vicinal 1H-1H coupling constants, NOE correlations, and by application of Kishi's universal NMR database. In order to get insights into the biosynthetic pathway of 1-3, the genome sequence of the producer strain D. aurantiacum was obtained and the putative biosynthetic gene cluster encoding their biosynthesis was identified through bioinformatic analysis using antiSMASH. Compounds 1-3 showed significant in-vitro antimycobacterial and cytotoxic activity.

Epoxynemanione A, nemanifuranones A–F, and nemanilactones A–C, from Nemania serpens, an endophytic fungus isolated from Riesling grapevines

Ten polyketide specialized metabolites, epoxynemanione A, nemanifuranones A–F, and nemanilactones A–C, were isolated from the culture filtrate of Nemania serpens (Pers.) Grey (1821), an endophytic fungus from a Riesling grapevine (Vitis vinifera) found in Canada's Niagara region. Additionally, four known metabolites 2-(hydroxymethyl)-3-methoxy-benzoic acid, phyllostine, 5-methylmellein and a nordammarane triterpenoid were isolated. A related known metabolite 2,3-dihydro-2-hydroxy-2,4-dimethyl-5-trans-propenylfuran-3-one has also been included for structural and biological comparison to the nemanifuranones. The latter was isolated from the culture filtrates of Mollisia nigrescens, an endophytic fungus from the leaves and stems of lowbush blueberry (Vaccinium angustifolium) found in the Acadian forest of Nova Scotia, Canada. Their structures were elucidated based on 1D and 2D NMR, HRESIMS measurements, X-ray crystallographic analysis of nemanifuranone A, the nordammarane triterpenoid and 2,3-dihydro-2-hydroxy-2,4-dimethyl-5-trans-propenylfuran-3-one compounds, and comparison of NOE and vicinal 1H-1H coupling constants to literature data for relative stereochemical assignments. Nemanifuranone A possesses a rare C2 hemiacetal and was active against both Gram-negative and Gram-positive bacteria.

JBIR-129 and -139, Cytotoxic 34-Membered Polyol Macrolides of Microbial Origin

New 34-membered polyol macrolides JBIR-129 (1) and JBIR-139 (2) were isolated from the culture of the terrestrial Streptomyces RK74. The planar structures of 1 and 2 were established on the basis of 1D and 2D NMR, ESI-TOF-MS, IR, and UV spectra. The relative configurations of the sugar units were determined by analyzing vicinal ¹H-¹H coupling constants and steric information. Both 1 and 2 showed cytotoxic activity against human ovarian adenocarcinoma SKOV-3 cells with IC₅₀ values of 0.3 and 0.4 μM, respectively.

Nonempirical study on the potential energy surface of 5-methyl-1,3-dioxane

Quantum-chemical study on the potential energy surface of 5-methyl-1,3-dioxane at the nonempirical RHF//STO-3G, RHF//3-21G, RHF//6-31G(d), RHF//6-31G(d,p), and MP2//6-31G(d,p) levels of theory revealed two energy-equivalent paths of conformational transformation of the equatorial and axial chair conformers. Potential barriers to these processes were estimated. The δG° value for the methyl substituent on C5 in 1,3-dioxane ring, determined on the basis of the experimental (NMR) and calculated vicinal 1H-1H coupling constants, was very consistent with published data

Characterization of the selenotrisulfide formed by reaction of selenite with end-capped phytochelatin-2.

The phytochelatins are a family of peptides synthesized by plants in response to exposure to heavy metals and metalloids, including selenium in the form of selenite. The amino acid sequence of the phytochelatin (PC) peptides is (gamma-Glu-Cys)n-Gly, where n typically ranges from 2 to 5. In this paper, the products of the reaction of selenite with an end-capped analogue of PC2, Ac-(gamma-Glu-Cys)2-Gly-NH2, are characterized. Selenite reacts with Ac-(gamma-Glu-Cys)2-Gly-NH2 (Ac-PC2-NH2) to form a compound that contains an intramolecular selenotrisulfide (-SSeS-)-linkage (Se[Ac-PC2-NH2]) and oxidized Ac-PC2-NH2. Both Se[Ac-PC2-NH2] and oxidized Ac-PC2-NH2 were isolated by HPLC and were characterized by MALDI-TOF mass spectrometry, by two-dimensional 1H and 13C NMR and, in the case of Se[Ac-PC2-NH2], by 77Se NMR. Using dihedral angles determined from vicinal 1H-1H coupling constants as constraints for the conformations around the Cys(CalphaH)-Cys(CbetaH) bonds, structures were predicted for the most abundant form of both compounds by Monte Carlo molecular mechanics simulations.

NMR study of the conformation of galactocerebroside in bilayers and solution: galactose reorientation during the metastable-stable gel transition.

Conformations of two types of bovine brain cerebroside containing normal and alpha-hydroxy-fatty acids (NFA-CER and HFA-CER, respectively) in solution and in bilayers were investigated using 1H and 13C NMR in solution and in the solid state. The analysis of vicinal 1H-1H coupling constants and NOE measurements in solution indicated that in both cerebrosides the predominant conformation about the O1-C1, C1-C2, and C2-C3 bonds is ap/-sc/ap, respectively. The remarkable similarity in the 13C NMR chemical shifts in solution and in hydrated liquid-crystalline bilayers indicated that both cerebrosides in bilayers assume conformation essentially identical to those in solution. The obtained 13C NMR spectra in solution were used as a reference for comparison with the variable-temperature 13C CP-MAS NMR spectra in the metastable and stable gel phases. The lack of chemical shift changes of polar carbon atoms upon cooling the HFA-CER bilayers below the Tm strongly suggests that the liquid-crystalline-metastable gel transition is not associated with a conformational change of the head group. The observed line broadening can be interpreted in terms of the hydrocarbon chain crystallization and slow dynamics of the head group in the metastable phase. On the other hand, the relaxation of the metastable gel phase of HFA-CER caused profound changes in the 13C spectra, primarily of the signals of the galactose C1, the ceramide C2, C4, and C5, and the carbonyl group. These changes are interpreted using the known dependence of the chemical shifts of anomeric carbon on the conformation about the O1-C1 bond to suggest that the gel phase relaxation involves a significant reorientation of the galactose moiety caused by a change in the rotation of the O1-C1 bond from the ap to -sc conformer. Similar changes of chemical shifts were observed in the case of NFA-CER during the transition from the liquid-crystalline phase to the stable gel phase.

A Practical Method for Stereospecific Assignments of γ- and δ-Methylene Hydrogens via Estimation of Vicinal 1H-1H Coupling Constants

Stereospecific assignments are made for γ- and δ-methylene hydrogens in a protein by means of estimation of vicinal 1H-1H spin-spin coupling constants from a short-mixing-time TOCSY experiment. 3Jαβ coupling constants, as measured from a P.E. COSY map, are shown to be well correlated with α-β cross-peak intensities of a short-mixing-time (10 ms) TOCSY map. The procedure is illustrated by application to a trypsin-inhibitor protein (Mr 7 Kd). Thus, γ-methylene hydrogens of isoleucine residues have been stereospecifically assigned on the basis of 3Jβγ1H-1H coupling patterns and intraresidue cross-peak intensities in a NOESY map; γ-hydrogens of other residues, such as lysine and arginine, have been stereospecifically assigned solely on the basis of cross-peak intensity patterns resulting from coupling of two β-hydrogens to two γ-hydrogens, and in conjunction with stereospecific assignments of β-methylene hydrogens. However, intraresidue NOE intensities are needed if one or two pairs of coupling constants cannot be estimated because of cross peaks either overlapping or occurring proximal to the diagonal. The δ-methylene hydrogens have been stereospecifically assigned on the basis of coupling between two γ-hydrogens and two δ-hydrogens, in combination with stereospecific assignments of γ-hydrogens. Stereospecific assignments of side chains should contribute to the overall precision and accuracy of NMR-determined three-dimensional solution structures of proteins.

14-membered cyclic opioids related to dermorphin and their partially retro-inverso modified analogues. II. Preferred conformations in solution as studied by 1H-NMR spectroscopy.

The 1H-NMR studies were extensively carried out to elucidate preferred conformations of a series of 14-membered cyclic dermorphin analogues containing two phenylalanines at both the third and fourth positions, e.g., Tyr-c[D-A2bu-Phe-Phe-(L and D)-Leu], Tyr-c[D-A2bu-Phe-gPhe-(S and R)-mLeu], and Tyr-c[D-Glu-Phe-gPhe-(L and D)-rLeu]. The temperature coefficients of the amide proton chemical shifts, vicinal 1H-1H coupling constants for the NH-CH groupings, and nuclear Overhauser effects provided information regarding the preferred conformations of the backbones. The conformational preferences and flexibility of the side chains were also estimated from the vicinal 1H-1H coupling constants around the C-C beta and C beta-C bonds in the articulated side chains. A comparison of the results obtained was made with the results previously obtained for the corresponding enkephalin analogues containing a glycine at the third position. It was found that the replacement of the glycine with the phenylalanine at the third position increases the conformational flexibility of the molecules with an L-, or S-, residue at the fifth position but reduces the flexibility of the molecules with D-, or R-, residue at the same position. The rotating frame nuclear Overhauser experiments gave direct evidence for compact conformations, with the Tyr side chain folding back over the 14-membered ring in Tyr-c[D-Glu-Phe-gPhe-rLeu], which displays relatively high selectivity for the delta-receptor over the mu-receptor. This observation is in agreement with our model proposed for the cyclic enkephalin analogues: folded forms with close aromatic ring placement are required for the activity at the delta-receptor.

Assessment of competing 2′«5′ versus 3′«5′ stackings in solution structure of branched-RNA by 1H- &31 P-NMR spectroscopy

Preparation of five novel phosphorylated derivatives of adenosine, i.e. adenosine 2′,3′-bis(ethylphosphate) (11), adenosine 2′,3′-bis(phosphate (13), adenosine 2′,3′,5′-tris(ethylphosphate (15), adenosine 2′,5′-bis(ethylphospahte) (17), and adenosine 3′,5′-bis(ethylphospahte) (19) is reported. These compounds, along with methyl β-D-ribofuranosyl-bis-2′,3′-ethylphosphate (9), were used as reference systems for 31P and1H-NMR conformational studies on the branched RNA structures 20 – 30. Compounds 11, 13, 15, 17, and 19 preserve the essential structural elements of the branch point adenosine, while the intramolecular base stacking interactions are removed. The 31P-NMR chemical shifts of 20 – 30, referenced against 9, 11, 13, or 15, show a pattern that is generally consistent with our previous results from variable temperture 31P-NMR experiments. The data indicate that the contribution of g,g around the P-O3′ (ζ) and P-05′ (α) boonds is significantly greater for the 2′-phosphate group than for the 3′-phosphate grooup. These results point towards preferential 2′→5′ rather than 3′→5′ base stacked structures in all of these synthetic models of the lariat. This is especially the case for the branched trimer 20 and the pentamer 27 which are part of a naturally occurring lariat structure. Note that the strongest 2′ →5′ stacking is however found in the unnatural trimers 22 and 23 in which the 2′-linked residue is a pyrimidine nucleotide. Compounds 11, 13, and 15 were also used to calibrate the 1H-NMR oligomerization shifts of the H2 protons of the branch-point adenosine. These data show a consistency with the results from variable temperature 1H-NMR experiments, as well as with the results of31P-NMR experiments. The results obtained with the series of compounds 20 (2′p5′G3′p5′U), 26a (UA2′p5′G3'o5′U, 27 (A2′p5′GU3′p5′UC), 28 (CUA2′p5′GU3′p5′UC), 29 (CUA2′p5′GUG3′p5′UCA), and 30 (CCUA2′p5′GUG3′p5′UCA) are of special interest since these structures are constituents of the naturally occurring lariat in the excised intron in Group II splicing of bl 1 of Yeast mitochondria. Qualitatively, the present 1H- and 31P-NMR data on 26a, 27, 28, 29, and 30 show 2′→5′ base stacking of an intermediate strength; 2′→5′ base stacking is substantially stronger for trimer 20 and pentamer 27. This difference is ascribed to the 5′-conformational transmission effect owing to the presence of at least one nucleotide upstream of the branch-point. 5′-Conformational transmission appears to weaken the 2′→5′ stacking at the expense of some 3′→5′ stacking. The experimental data on the conformation of 20 (A2′p5′g3′p5′U) (31 P and 1H chemical shifts, vicinal 1H-1H, 1H-31P, and 13C-31P coupling constants) formed the basis for a series of AMBER molecular mechanics calculations. These molecular modelling studies allowed us to conclude that g,g, conformation in the 2′-phosphate group is primarily g-(ζ),g- (α). This is found to be the only conformation that gives 2′→5′ base stacking as evident in the temperature dependent chemical shift and in the oligomerization shift studies. Modelling studies furthermore showed two energetically possible ζ and α torsions for the 3′-phosphate group (g−(ζ),g−(α) and g−(ζ),???(α)). The present use of reference compounds 9, 11, 13, 15, 17, and 19 has led to a refined and partially revised concept for the conformational description of oligomeric branched RNAs.

Significant conformational changes in an antigenic carbohydrate epitope upon binding to a monoclonal antibody

Transferred nuclear Overhauser enhancement spectroscopy (TRNOE) was used to observe changes in a ligand's conformation upon binding to its specific antibody. The ligands studied were methyl O-beta-D-galactopyranosyl(1----6)-4-deoxy-4-fluoro-beta-D-galactopyra nos ide (me4FGal2) and its selectively deuteriated analogue, methyl O-beta-D-galactopyranosyl(1----6)-4-deoxy-2-deuterio-4-fluoro-beta -D- galactopyranoside (me4F2dGal2). The monoclonal antibody was mouse IgA X24. The solution conformation of the free ligand me4F2dGal2 was inferred from measurements of vicinal 1H-1H coupling constants, long-range 1H-13C coupling constants, and NOE cross-peak intensities. For free ligand, both galactosyl residues adopt a regular chair conformation, but the NMR spectra are incompatible with a single unique conformation of the glycosidic linkage. Analysis of 1H-1H and 1H-13C constants indicates that the major conformer has an extended conformation: phi = -120 degrees; psi = 180 degrees; and omega = 75 degrees. TRNOE measurements on me4FGal2 and me4F2dGal2 in the presence of the specific antibody indicate that the pyranose ring pucker of each galactose ring remains unchanged, but rotations about the glycosidic linkage occur upon binding to X24. Computer calculations indicate that there are two sets of torsion angles that satisfy the observed NMR constraints, namely, phi = -152 +/- 9 degrees; psi = -128 +/- 7 degrees; and omega = -158 +/- 6 degrees; and a conformer with phi = -53 +/- 6 degrees; psi = 154 +/- 10 degrees; and omega = -173 +/- 6 degrees. Neither conformation is similar to any of the observed conformations of the free disaccharide.(ABSTRACT TRUNCATED AT 250 WORDS)

13C and 1H NMR spectra of all methyl O-benzoyl-β-D-xylopyranosides. nonadditivity of acylation effects on 13C chemical shifts in deutoriochloroform solutions

All methyl O-benzoyl-β-D-xylopyranosides have been prepared and their 1H and 13C NMR spectra measured in deuteriochloroform solutions. The 1H NMR spectra were analysed to the first order and assigned with the aid of homonuclear decoupling. The 13C chemical shifts were assigned through heteronuclear selective decouling experiments. Some of the 13C chemical shifts observed in di- and tri-O-benzoyl derivatives differ considerably from those calculated according to the direct additivity rule from the shifts in the mono derivatives. It is shown that the nonadditivity is due to a conformational heterogeneity of the series of investigated compounds dissolved in deuteriochloroform. The heterogeneity is evidenced by the vicinal 1H-1H coupling constants and by 13 chemical shifts of C(1) methoxyl carbon atoms.

Conformational studies of some 2':3'-cyclic mononucleotides in solution by different nuclear-magnetic-resonance methods.

The conformations of the 2':3'-cyclic mononucleotides of adenosine and cytidine in deuterium oxide has been studied at pH 2.3, using lanthanide ions as paramagnetic nuclear magnetic resonance (NMR) probes. It was not possible to find any single conformation for these molecules which accounts for the observed shift and relaxation data. This situation is in agreement with the interpretation of vicinal 1H-1H and 1H-31P coupling constants, which indicate that the ribofuranose and cyclic phosphate rings are in rapid equilibrium between different puckered forms. The interpretation of the lanthanide data in terms of an equilibrium between different conformations give average rotamer populations in good agreement with the coupling constant analysis. The conformations of these systems in aqueous solutions were found to be more flexible than in the solid state, where rigid planar ribofuranose rings have been observed. Adenosine 2':3'-monophosphate differs from cytidine 2':3'-monophosphate at the glycosidic link.