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1.
Boisbouvier J Bryce DL O'neil-Cabello E Nikonowicz EP Bax A 《Journal of biomolecular NMR》2004,30(3):287-301
New methods are described for accurate measurement of multiple residual dipolar couplings in nucleic acid bases. The methods use TROSY-type pulse sequences for optimizing resolution and sensitivity, and rely on the E.COSY principle to measure the relatively small two-bond 2DCH couplings at high precision. Measurements are demonstrated for a 24-nt stem-loop RNA sequence, uniformly enriched in 13C, and aligned in Pf1. The recently described pseudo-3D method is used to provide homonuclear 1H-1H decoupling, which minimizes cross-correlation effects and optimizes resolution. Up to seven 1H-13C and 13C-13C couplings are measured for pyrimidines (U and C), including 1DC5H5, 1DC6H6, 2DC5H6, 2DC6H5, 1DC5C4, 1DC5C6, and 2DC4H5. For adenine, four base couplings (1DC2H2, 1DC8H8, 1DC4C5, and 1DC5C6) are readily measured whereas for guanine only three couplings are accessible at high relative accuracy (1DC8H8, 1DC4C5, and 1DC5C6). Only three dipolar couplings are linearly independent in planar structures such as nucleic acid bases, permitting cross validation of the data and evaluation of their accuracies. For the vast majority of dipolar couplings, the error is found to be less than ±3% of their possible range, indicating that the measurement accuracy is not limiting when using these couplings as restraints in structure calculations. Reported isotropic values of the one- and two-bond J couplings cluster very tightly for each type of nucleotide. 相似文献
2.
Ming Tang Gemma Comellas Leonard J. Mueller Chad M. Rienstra 《Journal of biomolecular NMR》2010,48(2):103-111
High resolution 13C-detected solid-state NMR spectra of the deuterated beta-1 immunoglobulin binding domain of the protein G (GB1) have been
collected to show that all 15N, 13C′, 13Cα and 13Cβ sites are resolved in 13C–13C and 15N–13C spectra, with significant improvement in T
2 relaxation times and resolution at high magnetic field (750 MHz). The comparison of echo T
2 values between deuterated and protonated GB1 at various spinning rates and under different decoupling schemes indicates that
13Cα T
2′ times increase by almost a factor of two upon deuteration at all spinning rates and under moderate decoupling strength,
and thus the deuteration enables application of scalar-based correlation experiments that are challenging from the standpoint
of transverse relaxation, with moderate proton decoupling. Additionally, deuteration in large proteins is a useful strategy
to selectively detect polar residues that are often important for protein function and protein–protein interactions. 相似文献
3.
An experiment is presented to determine 3JHNHα coupling constants, with significant advantages for applications to unfolded proteins. The determination of coupling constants
for the peptide chain using 1D 1H, or 2D and 3D 1H-15N correlation spectroscopy is often hampered by extensive resonance overlap when dealing with flexible, disordered proteins.
In the experiment detailed here, the overlap problem is largely circumvented by recording 1H-13C′ correlation spectra, which demonstrate superior resolution for unfolded proteins. J-coupling constants are extracted from
the peak intensities in a pair of 2D spin-echo difference experiments, affording rapid acquisition of the coupling data. In
an application to the cytoplasmic domain of human neuroligin-3 (hNlg3cyt) data were obtained for 78 residues, compared to 54 coupling constants obtained from a 3D HNHA experiment. The coupling constants
suggest that hNlg3cyt is intrinsically disordered, with little propensity for structure. 相似文献
4.
One bond methyl 1H-13C and 13Cmethyl–13C scalar and residual dipolar couplings have been measured at sites in an 15N, 13C, 50% 2H labeled sample of the B1 immunoglobulin binding domain of peptostreptococcal protein L to investigate changes in the structure of methyl groups in response to deuterium substitution. Both one bond methyl 1H-13C and 13Cmethyl–13C scalar coupling constants have been found to decrease slightly with increasing deuterium content. Previous studies have shown that 1H-13C couplings in methyl groups are exquisitely sensitive to electronic structure, with decreases in coupling values as a function of deuteration consistent with a slight lengthening of the remaining H-C bonds. Changes in the HmethylCmethylC angle are found to be small, with average differences on the order of 0.3 ± 0.1° and 0.4 ± 0.2° between CH3, CH2D and CH3, CHD2 isotopomers, respectively. Knowledge of methyl geometry is a prerequisite for the extraction of accurate dynamics parameters from spin relaxation studies involving these groups. 相似文献
5.
《Phytochemistry》1986,25(3):751-752
Structure elucidation and total assignment of the 13C NMR spectrum of 12-(S)-hydroxygeranylgeraniol, a new acyclic diterpene from the grown alga Bifurcaria bifurcata, was accomplished through the use of 1HNMR, 13C NMR and 2D NMR spectroscope including 2D long range 1H-13C chemical shift correlations. 相似文献
6.
Solution structure of the DNA-binding domain of the heat shock transcription factor determined by multidimensional heteronuclear magnetic resonance spectroscopy
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Fred F. Damberger Jeffrey G. Pelton Celia J. Harrison Hillary C.M. Nelson David E. Wemmer 《Protein science : a publication of the Protein Society》1994,3(10):1806-1821
7.
Tommy ?man May-Britt Tessem Tone F Bathen Helena Bertilsson Anders Angelsen Mattias Hedenstr?m Trygve Andreassen 《BMC bioinformatics》2014,15(1)
Background
Identification of individual components in complex mixtures is an important and sometimes daunting task in several research areas like metabolomics and natural product studies. NMR spectroscopy is an excellent technique for analysis of mixtures of organic compounds and gives a detailed chemical fingerprint of most individual components above the detection limit. For the identification of individual metabolites in metabolomics, correlation or covariance between peaks in 1H NMR spectra has previously been successfully employed. Similar correlation of 2D 1H-13C Heteronuclear Single Quantum Correlation spectra was recently applied to investigate the structure of heparine. In this paper, we demonstrate how a similar approach can be used to identify metabolites in human biofluids (post-prostatic palpation urine).Results
From 50 1H-13C Heteronuclear Single Quantum Correlation spectra, 23 correlation plots resembling pure metabolites were constructed. The identities of these metabolites were confirmed by comparing the correlation plots with reported NMR data, mostly from the Human Metabolome Database.Conclusions
Correlation plots prepared by statistically correlating 1H-13C Heteronuclear Single Quantum Correlation spectra from human biofluids provide unambiguous identification of metabolites. The correlation plots highlight cross-peaks belonging to each individual compound, not limited by long-range magnetization transfer as conventional NMR experiments.Electronic supplementary material
The online version of this article (doi:10.1186/s12859-014-0413-z) contains supplementary material, which is available to authorized users. 相似文献8.
《Phytochemistry》1986,25(6):1453-1459
The aerial part of Narcissus requienii (Amaryllidaceae) was found to contain pseudolycorine as the major alkaloid, and two new phenolic bases: 2-O-acetylpseudolycorine and 1-O-acetylpseudolycorine. The present communication reports the complete assignments of the NMR spectra of pseudolycorine, and the structure elucidation of the other comnounds by means of 2D 1H-1H and 1H-13C NMR chemical shift correlation experiments. 相似文献
9.
Polyester Biosynthesis Characteristics of Pseudomonas citronellolis Grown on Various Carbon Sources, Including 3-Methyl-Branched Substrates
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Forty-two different carbon sources were tested for the polyester synthesis of a citronellol-utilizing bacterium, Pseudomonas citronellolis (ATCC 13674). These included linear C2 to C10 monocarboxylic acids, C3 to C10 dicarboxylic acids, saccharides, α,ω-diols, hydrocarbons, and 3-methyl-branched substrates such as 3,7-dimethyl-6-octen-1-ol (citronellol), 3-methyl-n-valerate, 3-methyl-1-butanol, and 3-methyladipate. Isolated polymers were characterized by gas chromatography, infrared spectroscopy, 1H- or 13C-nuclear magnetic resonance spectroscopy, 1H-13C heteronuclear correlation spectroscopy (1H-13C COSY), 1H-1H homonuclear COSY, and differential scanning calorimetry. Polyesters from nine monocarboxylic acids and two related carbon sources could be metabolically divided into three groups. The first group of C2 to C4 carbon sources resulted in copolyesters composed of 61 to 70 mol% 3-hydroxydecanoate, 23 to 33 mol% 3-hydroxyoctanoate, 3.6 to 9.0 mol% 3-hydroxy-5-cis-dodecenoate, and 1.8 to 2.6 mol% 3-hydroxy-7-cis-tetradecenoate. Carbon sources in group II (C7 to C10) produced copolyesters composed of 3-hydroxyacid monomer units with the same number of carbon atoms as the substrate (major constituent) and monomer units with either two less or two more carbons. Negligible amounts of 3-hydroxy-5-cis-dodecenoate and 3-hydroxy-7-cis-tetradecenoate were detected in copolyesters from this group. Copolyesters from group III (C5 and C6) had a monomer unit distribution that could be said to be between those of groups I and II. In addition, a novel copolyester, poly(3-hydroxy-7-methyl-6-octenoate-co-3-hydroxy-5-methylhexanoate), was synthesized when grown on citronellol. The 1H-13C heteronuclear COSY spectrum for monomer unit II revealed that both methylene and isopropyl groups, proximately connected in series to a single chiral center, had magnetically diastereotopic natures. 相似文献
10.
Aim: To purify and characterize an antimicrobial compound produced by a biocontrol bacterium, Pseudomonas aeruginosa MML2212, and evaluate its activity against rice pathogens, Rhizoctonia solani and Xanthomonas oryzae pv. oryzae. Methods and Results: Pseudomonas aeruginosa strain MML2212 isolated from the rice rhizosphere with wide‐spectrum antimicrobial activity was cultured in Kings’B broth using a fermentor for 36 h. The extracellular metabolites were isolated from the fermented broth using ethyl acetate extraction and purified by two‐step silica‐gel column chromatography. Three fractions were separated, of which a major compound was obtained in pure state as yellow needles. It was crystallized after dissolving with chloroform followed by slow evaporation. It is odourless with a melting point of 220–222°C. It was soluble in most of the organic solvents and poorly soluble in water. The molecular mass of purified compound was estimated as 223·3 by mass spectral analysis. Further, it was characterized by IR, 1H and 13C NMR spectral analyses. The crystal structure of the compound was elucidated for the first time by X‐ray diffraction study and deposited in the Cambridge Crystallographic Data Centre ( http://www.ccde.com.ac.uk ) with the accession no. CCDC 617344 . Conclusion: The crystal compound was undoubtedly identified as phenazine‐1‐carboxamide (PCN) with the empirical formula of C13H9N3O. Significance and Impact of the Study: As this is the first report on the crystal structure of PCN, it provides additional information to the structural chemistry. Furthermore, the present study reports the antimicrobial activity of purified PCN on major rice pathogens, R. solani and X. oryzae pv. oryzae. Therefore, the PCN can be developed as an ideal agrochemical candidate for the control of both sheath blight and bacterial leaf blight diseases of rice. 相似文献
11.
Summary Monodeuterated methylene positions exhibit substantially superior spectral characteristics in 1H–13C correlation experiments as compared to diprotio signals. A combination of 2H decoupling and multiplet editing of HMQC and HSQC experiments provides resolution enhancement for both stereoselective and random fractionally deuterated samples. For HMQC experiments with [2-2Hr, 2-13C]glycine-labeled E. coli thioredoxin (11.7 kDa), 3-fold increases in both 1H and 13C resolution result in a complementary 9-fold enhancement in sensitivity. Owing to a smaller improvement in 13C resolution, the corresponding enhancements for the HSQC experiment are 2-fold less. 相似文献
12.
Shingo Marumo Masato Katayama Eisaku Komori Yoko Ozaki Masahiro Natsume Satoru Kondo 《Bioscience, biotechnology, and biochemistry》2013,77(7):1967-1968
The effect of oryzalexin D, which has been isolated as a group of novel phytoalexins of rice plant, on DNA, RNA, protein, lipid and chitin biosyntheses, respiration and cell membrane permeability was investigated in Pyricularia oryzae. The concentration for 50% inhibition (ED50) by oryzalexin D of the mycelial growth of P. oryzae was 230 ppm. At this concentration, oryzalexin D inhibited equally the incorporation of [2–14C]thymidine, [2–14C]uridine, l-[U-14C]amino acid mixture, l-[methyl-14C]methionine and d-[l-14C]glucosamine into DNA, RNA, protein, lipid and chitin in intact cells, but did not inhibit these systems in a homogenate of the mycelia of P. oryzae. Oryzalexin D scarcely inhibited the respiration of the homogenate and mitochondria at ED50. On the other hand, oryzalexin D at ED50 caused leakage of potassium and inhibited the uptake of glutamate by mycelial cells of P. oryzae. These results suggest that interference with the cell membrane function is responsible for the primary mode of action.of oryzalexin D against P. oryzae. 相似文献
13.
Three new compounds, 17β-cevanin-6-oxo-5α,20β-diol yibeinine (1), 2-(tetrahydro-5-(2-hydroxyphenyl)-2H-pyran-3-yl) phenol (2), 1,3-O-diferuloyl-2-methoxypropane diol (3), as well as four known compounds (4–7), have been isolated from the ethanol extract of dried bulbs of Fritillaria pallidiflora Schrenk. All structures were determined based on their spectroscopic data (1D and 2D NMR (including 1H NMR, 13C NMR, 1H-1H COSY, HMBC, HSQC, HSQC-TOCSY, and NOESY experiments), and MS). Biological evaluation showed that compounds 1–4 inhibited the production of nitric oxide (NO) in LPS-stimulated RAW 264.7 cells with IC50 values of 18.0, 38.7, 29.5, and 47.1 μM, respectively. These results indicated that compound 1 has potential anti-inflammatory activity. 相似文献
14.
Siddhartha Sarma Russell J. DiGate Debra L. Banville R. D. Guiles 《Journal of biomolecular NMR》1996,8(2):171-183
Summary Modern multidimensional double- and triple-resonance NMR methods have been applied to assign the backbone and side-chain 13C resonances for both equilibrium conformers of the paramagnetic form of rat liver microsomal cytochrome b
5. The assignment of backbone 13C resonances was used to confirm previous 1H and 15N resonance assignments [Guiles, R.D. et al. (1993) Biochemistry, 32, 8329–8340]. On the basis of short- and medium-range NOEs and backbone 13C chemical shifts, the solution secondary structure of rat cytochrome b
5 has been determined. The striking similarity of backbone 13C resonances for both equilibrium forms strongly suggests that the secondary structures of the two isomers are virtually identical. It has been found that the 13C chemical shifts of both backbone and side-chain atoms are relatively insensitive to paramagnetic effects. The reliability of such methods in anisotropic paramagnetic systems, where large pseudocontact shifts can be observed, is evaluated through calculations of the magnitude of such shifts.Abbreviations DANTE
delays alternating with nutation for tailored excitation
- DEAE
diethylaminoethyl
- DQF-COSY
2D double-quantum-filtered correlation spectroscopy
- EDTA
ethylenediaminetetraacetic acid
- HCCH-TOCSY
3D proton-correlated carbon TOCSY experiment
- HMQC
2D heteronuclear multiple-quantum correlation spectroscopy
- HNCA
3D triple-resonance experiment correlating amide protons, amide nitrogens and alpha carbons
- HNCO
3D triple-resonance experiment correlating amide protons, amide nitrogens and carbonyl carbons
- HNCOCA
3D triple-resonance experiment correlating amide protons, amide nitrogens and alpha carbons via carbonyl carbons
- HOHAHA
2D homonuclear Hartmann-Hahn spectroscopy
- HOHAHA-HMQC
3D HOHAHA relayed HMQC
- HSQC
2D heteronuclear single-quantum correlation spectroscopy
- IPTG
isopropyl thiogalactoside
- NOESY
2D nuclear Overhauser enhancement spectroscopy
- NOESY-HSQC
3D NOESY relayed HSQC
- TOCSY
2D total correlation spectroscopy
- TPPI
time-proportional phase incrementation
- TSP
trimethyl silyl propionate 相似文献
15.
Vladimír Sklenář Robert D. Peterson Marita R. Rejante Juli Feigon 《Journal of biomolecular NMR》1993,3(6):721-727
Summary New 2D and 3D 1H-13C-15N triple resonance experiments are presented which allow unambiguous assignments of intranucleotide H1'-H8(H6) connectivities in 13C-and 15N-labeled RNA oligonucleotides. Two slightly different experiments employing double INEPT forward and back coherence transfers are optimized to obtain the H1'-C1'-N9/N1 and H8/H6-C8/C6-N9/N1 connectivities, respectively. The correlation of H1' protons to glycosidic nitrogens N9/N1 is obtained in a nonselective fashion. To correlate H8/H6 with their respective glycosidic nitrogens, selective 13C-refocusing and 15N-inversion pulses are applied to optimize the magnetization transfers along the desired pathway. The approach employs the heteronuclear one-bond spin-spin interactions and allows the 2D 1H-15N and 3D1H-13C-15N chemical shift correlation of nuclei along and adjacent to the glycosidic bond. Since the intranucleotide correlations obtained are based exclusively on through-bond scalar interactions, these experiments resolve the ambiguity of intra-and internucleotide H1'-H8(H6) assignments obtained from the 2D NOESY spectra. These experiments are applied to a 30-base RNA oligonucleotide which contains the binding site for Rev protein from HIV. 相似文献
16.
NMR experiments for the sign determination of homonuclear scalar and residual dipolar couplings 总被引:1,自引:0,他引:1
A modified version of the JHH-TOCSY experiment, `signed COSY', is presented that allows the determination of the sign of residual dipolar 1H-1H coupling constants with respect to the sign of one-bond 1H-X coupling constants in linear three-spin systems X-1H-1H, where X = 13C or 15N. In contrast to the original JHH-TOCSY experiments, the signs of J
HH couplings may be determined for CH2-CH2 moieties and for uniformly 13C/15N-labelled samples. In addition, sensitivity is enhanced, diagonal peaks are suppressed and cross peaks are observed only between directly coupled protons, as in a COSY spectrum. 相似文献
17.
Pengfei Zhang Yinwei You Yuan Song Youzhi Wang 《Biocontrol Science and Technology》2015,25(11):1285-1298
A novel entomopathogenic fungus of Locusta migratoria was identified as Aspergillus oryzae using a comparative sequence analysis of the internal transcribed spacer regions, aflatoxin B1 detection and morphological analysis. The fungus isolated from a dead locust collected in northwestern China was found to be pathogenic to the insect. Phylogenetic experiments revealed a 99% similarity between the fungus and those of three species, A. oryzae, Aspergillus flavus and Aspergillus parvisclerotigenus which are in the same branch of the Flavi section of the genus Aspergillus. Tests to detect aflatoxin B1 demonstrated that this fungus is a non-aflatoxin B1 producer, unlike A. parvisclerotigenus. Furthermore, morphological comparison with A. oryzae and A. flavus revealed that Aspergillus sp. XJ-1 belongs to A. oryzae, and named as A. oryzae XJ-1. The results of bioassays against third-instar locusts showed that mortality was dose-dependent and its median lethal concentrations were 3.3 × 108, 1.7 × 107 and 7.2 × 106 conidia/ml on the 10th-, 13th- and 15th-day post-inoculation. Therefore, the A. oryzae XJ-1 may have biocontrol potential against locusts. 相似文献
18.
The effect of deuteration on the 13C linewidths of U-13C, 15N 2D crystalline bacteriorhodopsin (bR) from Halobacterium salinarium, a 248-amino acid protein with seven-transmembrane (7TM) spanning regions, has been studied in purple membranes as a prelude to potential structural studies. Spectral doubling of resonances was observed for receptor expressed in 2H medium (for both 50:50% 1H:2H, and a more highly deuterated form) with the resonances being of similar intensities and separated by < 0.3 ppm in the methyl spectral regions in which they were readily distinguished. Line-widths of the methyl side chains were not significantly altered when the protein was expressed in highly deuterated medium compared to growth in fully protonated medium (spectral line widths were about 0.5 ppm on average for receptor expressed both in the fully protonated and highly deuterated media from the Cδ, Cγ1, and Cγ2 Ile 13C signals observed in the direct, 21-39 ppm, and indirect, 9-17 ppm, dimensions). The measured 13C NMR line-widths observed for both protonated and deuterated form of the receptor are sufficiently narrow, indicating that this crystalline protein morphology is suitable for structural studies. 1H decoupling comparison of the protonated and deuterated bR imply that deuteration may be advantageous for samples in which low power 1H decoupling is required. 相似文献
19.
Sequence-specific assignment of the backbone 1H-and 31P-NMR lines in a short DNA duplex with homo- and heteronuclear correlated spectroscopy 总被引:4,自引:0,他引:4
M. H. Frey W. Leupin O. W. Srensen W. A. Denny R. R. Ernst K. Wüthrich 《Biopolymers》1985,24(12):2371-2380
We report the assignment of the backbone 1H- and 31P-nmr lines in the synthetic hexadeoxyribonucleotide pentaphosphate duplex d(GCATGC)2, using double quantum filtered 1H-1H correlation spectroscopy, 1H observed 1H-31P heteronuclear correlation spectroscopy, and 31P relayed 1H-1H correlation spectroscopy. The strategy used enables one to make sequence-specific resonance assignments without reference to a known or assumed conformation of the DNA fragment. 相似文献
20.
Youlin Xia Paolo Rossi Marco Tonelli Chengdong Huang Charalampos G. Kalodimos Gianluigi Veglia 《Journal of biomolecular NMR》2017,69(1):45-52
TROSY-based triple resonance experiments are essential for protein backbone assignment of large biomolecular systems by solution NMR spectroscopy. In a survey of the current Bruker pulse sequence library for TROSY-based experiments we found that several sequences were plagued by artifacts that affect spectral quality and hamper data analysis. Specifically, these experiments produce sidebands in the 13C(t 1) dimension with inverted phase corresponding to 1HN resonance frequencies, with approximately 5% intensity of the parent 13C crosspeaks. These artifacts originate from the modulation of the 1HN frequency onto the resonance frequency of 13Cα and/or 13Cβ and are due to 180° pulses imperfections used for 1H decoupling during the 13C(t 1) evolution period. These sidebands can become severe for CAi, CAi?1 and/or CBi, CBi?1 correlation experiments such as TROSY-HNCACB. Here, we implement three alternative decoupling strategies that suppress these artifacts and, depending on the scheme employed, boost the sensitivity up to 14% on Bruker spectrometers. A class of comparable Agilent/Varian pulse sequences that use WALTZ16 1H decoupling can also be improved by this method resulting in up to 60–80% increase in sensitivity. 相似文献