共查询到20条相似文献,搜索用时 31 毫秒
1.
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
2.
Christian Herbst Jirada Herbst Michela Carella Jörg Leppert Oliver Ohlenschläger Matthias Görlach Ramadurai Ramachandran 《Journal of biomolecular NMR》2010,47(1):7-17
An approach for generating efficient RNnnS, nk {\rm{RN}}_{n}^{\nu_{\rm{S}}, {\nu_{\rm{k}}}} symmetry-based dual channel RF pulse schemes for γ-encoded broadband 15N–13C dipolar recoupling at high magic angle spinning frequencies is presented. The method involves the numerical optimisation
of the RF phase-modulation profile of the basic “R” element so as to obtain heteronuclear double quantum dipolar recoupling sequences with satisfactory magnetisation transfer
characteristics. The basic “R” element was implemented as a sandwich of a small number of short pulses of equal duration with each pulse characterised
by a RF phase and amplitude values. The performance characteristics of the sequences were evaluated via numerical simulations
and 15N–13C chemical shift correlation experiments. Employing such 13C–15N double-quantum recoupling sequences and the multiple receiver capabilities available in the current generation of NMR spectrometers,
the possibility to simultaneously acquire 3D NCC and CNH chemical shift correlation spectra is also demonstrated. 相似文献
3.
Mireia Güell Josep M. Luis Miquel Solà Per E. M. Siegbahn 《Journal of biological inorganic chemistry》2009,14(2):229-242
Tyrosinase catalyzes the ortho hydroxylation of monophenols and the subsequent oxidation of the diphenolic products to the resulting quinones. In efforts
to create biomimetic copper complexes that can oxidize C–H bonds, Stack and coworkers recently reported a synthetic μ-η2:η2-peroxodicopper(II)(DBED)2 complex (DBED is N,N′-di-tert-butylethylenediamine), which rapidly hydroxylates phenolates. A reactive intermediate consistent with a bis-μ-oxo-dicopper(III)-phenolate
complex, with the O–O bond fully cleaved, is observed experimentally. Overall, the evidence for sequential O–O bond cleavage
and C–O bond formation in this synthetic complex suggests an alternative mechanism to the concerted or late-stage O–O bond
scission generally accepted for the phenol hydroxylation reaction performed by tyrosinase. In this work, the reaction mechanism
of this peroxodicopper(II) complex was studied with hybrid density functional methods by replacing DBED in the μ-η2:η2-peroxodicopper(II)(DBED)2 complex by N,N′-dimethylethylenediamine ligands to reduce the computational costs. The reaction mechanism obtained is compared with the
existing proposals for the catalytic ortho hydroxylation of monophenol and the subsequent oxidation of the diphenolic product to the resulting quinone with the aim
of gaining some understanding about the copper-promoted oxidation processes mediated by 2:1 Cu(I)O2-derived species.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
4.
Recently, the prenyltransferase SirD was found to be responsible for the O-prenylation of tyrosine in the biosynthesis of sirodesmin PL in Leptosphaeria maculans. In this study, the behavior of SirD towards phenylalanine/tyrosine and tryptophan derivatives was investigated. Product
formation has been observed with 12 of 19 phenylalanine/tyrosine derivatives. It was shown that the alanine structure attached
to the benzene ring and an electron donor, e.g., OH or NH2, at its para-position are essential for the enzyme activity. Modifications were possible both at the side chain and the benzene ring.
Enzyme products from seven phenylalanine/tyrosine derivatives were isolated and characterized by MS and NMR analyses including
HSQC and HMBC and proven to be O- or N-prenylated derivatives at position C4 of the benzene rings. K
M
values of six selected derivatives were found in the range of 0.10–0.68 mM. Catalytic efficiencies (K
cat/K
M
) were determined in the range of 430–1,110 s−1·M−1 with l-tyrosine as the best substrate. In addition, 7 of 14 tested tryptophan analogs were also accepted by SirD and converted to
C7-prenylated derivatives, which was confirmed by comparison with products obtained from enzyme assays using a 7-dimethylallyltryptophan
synthase 7-DMATS from Aspergillus fumigatus. 相似文献
5.
J Xu L Yang SJ Zhao ZT Wang ZB Hu 《World journal of microbiology & biotechnology》2012,28(4):1803-1806
Biotransformation of naringenin with Aspergillus niger CGMCC 3.4628 yielded two hydroxylation products which were identified unambiguously as 6-hydroxylnaringenin (carthamidin)
and 8-hydroxylnaringenin (isocarthamidin) by ESI–MS and 1H-NMR. Both products simultaneously arrived at high level after 48 h in the biotransformation process. The highest conversion
efficiency of carthamidin was 0.38 mg/mg of naringenin and that of isocarthamidin was 0.43 mg/mg of naringenin. Antioxidant
property assay using a thin layer chromatography-bioautographic-based DPPH scavenging method demonstrated that both hydroxylation
metabolites exhibited much stronger activity than naringenin. The high efficiency and convenient procedure makes the biotransformation
with A. niger described in current work a potential way to produce carthamidin and isocarthamidin. 相似文献
6.
James S. Craigie Shawna L. MacKinnon John A. Walter 《Journal of applied phycology》2008,20(5):665-671
Aqueous extracts of Ascophyllum nodosum and several other brown seaweeds are manufactured commercially and widely distributed for use on agricultural crops. The
increasingly regulated international trade in such products requires that they be standardized and defined to a degree not
previously required. We examined commercially available extracts using quantitative 1H NMR and principal components analysis (PCA) techniques. Extracts manufactured over a 4-year period using the same process
exhibited characteristic profiles that, on PCA, clustered as a discrete group distinct from the other commercial products
examined. In addition to recognizing extracts made from different seaweeds, analysis of the 1H spectra in the 0.35–4.70 ppm region allowed us to distinguish amongst extracts produced from the same algal species by different
manufacturers. This result established that the process used to make an extract is an important variable in defining its composition.
A comparison of the 1H NMR integrals for the regions 1.0–3.0 ppm and 3.0–4.38 ppm revealed small but significant changes in the A. nodosum spectra that we attribute to seasonal variation in gross composition of the harvested seaweed. Such changes are reflected
in the PCA scores plots and contribute to the scatter observed within the data point cluster observed for Acadian soluble
extracts when all data are pooled. Quantitative analysis using 1H NMR (qNMR) with a certified external standard (caffeine) showed a linear relationship with extract concentration over at
least an order of magnitude (2.5–33 mg/mL; R
2 > 0.97) for both spectral regions integrated. We conclude that qNMR can be used to profile (or “fingerprint”) commercial
seaweed extracts and to quantify the amount of extract present relative to a suitably chosen standard.
Issued as NRCC no. 42,652. 相似文献
7.
The surface dynamics of bacteriorhodopsin was examined by measurements of site-specific 13C–1H dipolar couplings in [3-13C]Ala-labeled bacteriorhodopsin. Motions of slow or intermediate frequency (correlation time <50 µs) scale down 13C–1H dipolar couplings according to the motional amplitude. The two-dimensional dipolar and chemical shift (DIPSHIFT) correlation technique was utilized to obtain the dipolar coupling strength for each resolved peak in the 13C MAS solid-state NMR spectrum, providing the molecular order parameter of the respective site. In addition to the rotation of the Ala methyl group, which scales the dipolar coupling to 1/3 of the rigid limit value, fluctuations of the C–C vector result in additional motional averaging. Typical order parameters measured for mobile sites in bacteriorhodopsin are between 0.25 and 0.29. These can be assigned to Ala103 of the C–D loop and Ala235 at the C-terminal -helix protruded from the membrane surface, and Ala196 of the F–G loop, as well as to Ala228 and Ala233 of the C-terminal -helix and Ala51 from the transmembrane -helix. Such order parameters departing significantly from the value of 0.33 for rotating methyl groups are obviously direct evidence for the presence of fluctuation motions of the Ala C–C vectors of intact preparations of fully hydrated, wild-type bacteriorhodopsin at ambient temperature. The order parameter for Ala160 from the expectantly more flexible E–F loop, however, is unavailable under highest-field NMR conditions, probably because increased chemical shift anisotropy together with intrinsic fluctuation motions result in an unresolved 13C NMR signal. 相似文献
8.
Valerio Pieri Sonja Sturm Christoph Seger Chlodwig Franz Hermann Stuppner 《Metabolomics : Official journal of the Metabolomic Society》2012,8(2):335-346
The characterization of T. vulgaris plant material for quality control purposes was performed by NMR-based methods. Direct extraction of 141 T. vulgaris samples with DMSO-d
6 enabled the obtainment of crude extracts with a representative composition in terms of both volatile and non-volatile constituents.
The acquisition of 600 MHz 1H NMR spectra resulted in a dataset which was analyzed by a combination of metabolic profiling and target analysis approaches.
Preliminary analysis of the 1H NMR spectra was performed by principal component analysis, which revealed sample discrimination on a chemotype basis (thymol,
carvacrol and linalool chemotypes). Further minor discriminative constituents were identified as p-cymene, γ-terpinene, rosmarinic acid, and 3,4,3′,4′-tetrahydroxy-5,5′-diisopropyl-2,2′-dimethylbiphenyl. Metabolite identification
was accomplished by 1D and 2D NMR techniques and supported by spiking experiments. Fast dereplication of constituents not
available as reference compounds was performed by HPLC–SPE–NMR experiments. A targeted approach based on qHNMR was validated
for quantification of the identified secondary metabolites. Validation was performed in terms of precision (intra-day RSD ≤ 4.51%,
inter-day RSD ≤ 4.18%), repeatability (RSD ≤ 2.30%), accuracy (recovery rates within 93.4 and 103.4%), linearity (correlation
coefficients ≥ 0.9990), robustness, and stability. The amount of the dominant monoterpene in thymol, carvacrol, and linalool
chemotypes was respectively found to be within 0.4–2.6, 0.7–2.3, and 1.1–3.6% (w/w). Variable amounts of the precursors p-cymene and γ-terpinene were found in thymol and carvacrol chemotypes. The highest amount of rosmarinic acid and 3,4,3′,4′-tetrahydroxy-5,5′-diisopropyl-2,2′-dimethylbiphenyl
in the analyzed samples was respectively 4.6 and 0.4% (w/w). Since quantification is performed on a weight basis, the essential
oil content can be estimated based on the sum of the quantified monoterpenes. The NMR-based analysis of T. vulgaris represents a more comprehensive approach in comparison to traditional chromatographic methods such as GC and LC, respectively
employed for the analysis of volatile and non-volatile constituents. Further advantages lie in the simple sample preparation,
rapidity and reproducibility of the NMR analysis. 相似文献
9.
Wild-type cmFDH contains no cystines, hence it is a good candidate to test the hypothesis that thermostability can be achieved by introducing
new disulphide bridges. Three cysteine double mutants of cmFDH were designed, using a homology model reported previously, to introduce cystine bridges in the C-domain (T169C–T226C)
in the N-domain (V88C–V112C) and between the two monomers (M156C–L159C) to form two cystine bridges across the dimer interface.
These mutants were constructed and the proteins were over-expressed in E. coli. The mutants V88C–V112C and M156C–L159C lost FDH activity. The mutant T169C–T226C was both less active and less thermostable
than wild-type FDH. 相似文献
10.
Xuesong Zhao Juan Wang Jie Li Ling Fu Juan Gao Xiuli Du Hongtao Bi Yifa Zhou Guihua Tai 《Journal of industrial microbiology & biotechnology》2009,36(5):721-726
Fourteen phytopathogenic fungi were tested for their ability to transform the major ginsenosides to the active minor ginsenoside
Rd. The transformation products were identified by TLC and HPLC, and their structures were assigned by NMR analysis. Cladosporium fulvum, a tomato pathogen, was found to transform major ginsenoside Rb1 to Rd as the sole product. The following optimum conditions for transforming Rd by C. fulvum were determined: the time of substrate addition, 24 h; substrate concentration, 0.25 mg ml−1; temperature, 37°C; pH 5.0; and biotransformation period, 8 days. At these optimum conditions, the maximum yield was 86%
(molar ratio). Further, a preparative scale transformation with C. fulvum was performed at a dose of 100 mg of Rb1 by a yield of 80%. This fungus has potential to be applied on the preparation for Rd in pharmaceutical industry. 相似文献
11.
Ulrich Metzger Ulrich Lankes Kai Fischpera Fritz H. Frimmel 《Applied microbiology and biotechnology》2009,85(1):197-206
Extracellular polymeric substances were extracted from the bacterial strain Pseudomonas putida and the fungal species Aureobasidium pullulans using three different methods (formaldehyde–NaOH, ethylenediaminetetraacetic acid (EDTA) and cation-exchange-resin). The
composition of the extracellular polymeric substances (EPS) was analysed by biochemical and high-resolution solid state 13C nuclear magnetic resonance (NMR) spectroscopic methods. The EPS yield was strongly dependent on the extraction method, with
the formaldehyde–NaOH method showing the best extraction efficiency. The NMR method revealed that when using the EDTA extraction
method, about 40% of the EDTA accumulated in the EPS and that was responsible for the apparent high extraction yields. EPS
protein content determined by the NMR method was up to 30% higher than the protein content determined using the biochemical
(Lowry) method for P. putida and for A. pullulans. The average protein carbon content determined by the NMR method was approximately 70% of the total carbon content. NMR results
could be supported by elemental analysis, which showed a high nitrogen content (~10%) in the EPS. The carbohydrate carbon
content detected with both methods in the cell aggregates and the EPS was approximately 20% in each. In this study, quantitative
13C cross-polarisation magic angle spinning NMR spectroscopy was conducted on unlabeled cell strains, and EPS and could be used
to quantify protein and carbohydrate of different samples. 相似文献
12.
We have developed NMR spectroscopic methods to investigate the tyrosines within Bacillus circulans xylanase (BcX). Four slowly exchanging buried tyrosine hydroxyl protons with chemical shifts between 7.5 and 12.5 ppm were
found using a long-range 13C-HSQC experiment that exploits the 3JCH coupling between the ring 1Hη and 13Cε nuclei. The NMR signals from these protons were assigned via 13C-tyrosine selective labelling and a suite of scalar and 13C,15N-filtered/edited NOE correlation spectra. Of the fifteen tyrosines in BcX, only the buried Tyr79 and Tyr105 showed four distinct,
rather than two averaged, signals from ring 13C–1H pairs, indicative of slow flipping on the chemical shift timescale. Ring flipping rate constants of ~10 and ~0.2 s−1 were measured for the two residues, respectively, using a 13C longitudinal exchange experiment. The hydrogen bonding properties of the Tyr79 and Tyr105 hydroxyls were also defined by
complementary NOE and J-coupling measurements. The 1Hη hydrogen–deuterium exchange rate constants of the buried tyrosines were determined from 13C/15N-filtered spectra recorded as a function of pH. These exchange rate constants correspond to estimated protection factors
of ~104–108 relative to a random coil tyrosine. The phenolic sidechain pK
a values were also measured by monitoring their pH-dependent 13Cζ chemical shifts via 1Hε/δ(13Cε)13Cζ correlation spectra. Exposed tyrosines had unperturbed pK
a values of ~10.2, whereas buried residues remained predominantly neutral at or even above pH 11. Combined with selective isotope
labelling, these NMR experiments should prove useful for investigating the structural and electrostatic properties of tyrosines
in many interesting proteins. 相似文献
13.
Carbonyl 13C′ relaxation is dominated by the contribution from the 13C′ chemical shift anisotropy (CSA). The relaxation rates provide useful and non-redundant structural information in addition
to dynamic parameters. It is straightforward to acquire, and offers complimentary structural information to the 15N relaxation data. Furthermore, the non-axial nature of the 13C′ CSA tensor results in a T1/T2 value that depends on an additional angular variable even when the diffusion tensor of the protein molecule is axially symmetric.
This dependence on an extra degree of freedom provides new geometrical information that is not available from the NH dipolar
relaxation. A protocol that incorporates such structural restraints into NMR structure calculation was developed within the
program Xplor-NIH. Its application was illustrated with the yeast Fis1 NMR structure. Refinement against the 13C′ T1/T2 improved the overall quality of the structure, as evaluated by cross-validation against the residual dipolar coupling as
well as the 15N relaxation data. In addition, possible variations of the CSA tensor were addressed.
Electronic Supplementary Material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
14.
Acaryochloris marina is an oxygen-evolving organism that utilizes chlorophyll-d for light induced photochemistry. In photosystem I particles from Acaryochloris marina, the primary electron donor is called P740, and it is thought that P740 consist of two chlorophyll-d molecules. (P740+-P740) FTIR difference spectra have been produced, and vibrational features that are specific to chlorophyll-d (and not chlorophyll-a) were observed, supporting the idea that P740 consists chlorophyll-d molecules. Although bands in the (P740+-P740) FTIR difference spectra were assigned specifically to chlorophyll-d, how these bands shifted, and how their intensities changed, upon cation formation was never considered. Without this information
it is difficult to draw unambiguous conclusions from the FTIR difference spectra. To gain a more detailed understanding of
cation induced shifting of bands associated with vibrational modes of P740 we have used density functional theory to calculate
the vibrational properties of a chlorophyll-d model in the neutral, cation and anion states. These calculations are shown to be of considerable use in interpreting bands
in (P740+-P740) FTIR difference spectra. Our calculations predict that the 31 formyl C–H mode of chlorophyll-d upshifts/downshifts upon cation/anion formation, respectively. The mode intensity also decreases/increases upon cation/anion
formation, respectively. The cation induced bandshift of the 31 formyl C–H mode of chlorophyll-d is also strongly dependant on the dielectric environment of the chlorophyll-d molecules. With this new knowledge we reassess the interpretation of bands that were assigned to 31 formyl C–H modes of chlorophyll-d in (P740+-P740) FTIR difference spectra. Considering our calculations in combination with (P740+-P740) FTIR DS we find that the most likely conclusions are that P740 is a dimeric Chl-d species, in an environment of low effective dielectric constant (∼2–8). In the P740+ state, charge is asymmetrically distributed over the two Chl-d pigments in a roughly 60:40 ratio.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
15.
The objective of this study was to improve the biological water–gas shift reaction for producing hydrogen (H2) by conversion of carbon monoxide (CO) using an anaerobic thermophilic pure strain, Carboxydothermus hydrogenoformans. Specific hydrogen production rates and yields were investigated at initial biomass densities varying from 5 to 20 mg volatile
suspended solid (VSS) L−1. Results showed that the gas–liquid mass transfer limits the CO conversion rate at high biomass concentrations. At 100-rpm
agitation and at CO partial pressure of 1 atm, the optimal substrate/biomass ratio must exceed 5 mol CO g−1 biomass VSS in order to avoid gas–liquid substrate transfer limitation. An average H2 yield of 94 ± 3% and a specific hydrogen production rate of ca. 3 mol g−1 VSS day−1 were obtained at initial biomass densities between 5 and 8 mg VSS−1. In addition, CO bioconversion kinetics was assessed at CO partial pressure from 0.16 to 2 atm, corresponding to a dissolved
CO concentration at 70°C from 0.09 to 1.1 mM. Specific bioactivity was maximal at 3.5 mol CO g−1 VSS day−1 for a dissolved CO concentration of 0.55 mM in the culture. This optimal concentration is higher than with most other hydrogenogenic
carboxydotrophic species. 相似文献
16.
Moslem Papizadeh Mohammad Roayaei Ardakani Gholamhossein Ebrahimipour Hossein Motamedi 《World journal of microbiology & biotechnology》2010,26(7):1195-1200
Oil-polluted soils were sampled from National Iranian South Oil Company (NISOC) for isolation and screening of C–S and not
C–C targeted Dibenzothiophene (DBT) degrading microorganisms. Microbacterium sp. NISOC-06, a C–S targeted DBT degrading bacterium, was selected and its desulfurization ability was studied in aqueous phase and water-gasoline
biphasic systems. The 16srRNA gene was amplified using universal eubacteria-specific primers, PCR product was sequenced and
the sequence of nearly 1,500 bp 16srDNA was studied. Based on Gas Chromatography results Microbacterium sp. NISOC-06 utilized 94.8% of 1 mM DBT during the 2 weeks of incubation. UV Spectrophotometry and biomass production measurements showed
that the Microbacterium sp. NISOC-06 was not able to utilize DBT as a carbon source. There was no accumulation of phenolic compounds as Gibb’s assay showed. Biomass
production in a biphasic system for which DBT-enriched gasoline was used as the sulfur source indicated the capability of
Microbacterium sp. NISOC-06 to desulfurize gasoline. 相似文献
17.
S Smaoui F Mathieu L Elleuch Y Coppel G Merlina I Karray-Rebai L Mellouli 《World journal of microbiology & biotechnology》2012,28(3):793-804
A new actinomycete strain designated TN256, producing antimicrobial activity against pathogenic bacteria and fungi, was isolated
from a Tunisian Saharan soil. Morphological and chemical studies indicated that strain TN256 belonged to the genus Streptomyces. Analysis of the 16S rDNA sequence of strain TN256 showed a similarity level ranging between 99.79 and 97.8% within Streptomyces microflavus DSM 40331T and Streptomyces griseorubiginosus DSM 40469T respectively. The comparison of its physiological characteristics showed significant differences with the nearest species.
Combined analysis of the 16 S rRNA gene sequences (FN687758), fatty acids profile, and results of physiological and biochemical
tests indicated that there were genotypic and phenotypic differentiations of that isolate from other Streptomyces species neighbours. These date strongly suggest that strain TN256 represents a novel species with the type strain Streptomyces TN256 (=CTM50228T). Experimental validation by DNA–DNA hybridization would be required for conclusive confirmation. Four active products (1–4)
were isolated from the culture broth of Streptomyces TN256 using various separation and purification steps and procedures. 1: N-[2-(1H-indol-3-yl)-2 oxo-ethyl] acetamide ‘alkaloid’ derivative; 2: di-(2-ethylhexyl) phthalate, a phthalate derivative; 3: 1-Nonadecene and 4: Cyclo (l-Pro-l-Tyr) a diketopiperazine ‘DKP’ derivative. The chemical structure of these four active compounds was established on the basis
of spectroscopic studies NMR and by comparing with data from the literature. According to our biological studies, we showed
in this work that the pure compounds (1–4) possess antibacterial and antifungal activities. 相似文献
18.
Engineering <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis> for the production of pyruvate 总被引:1,自引:0,他引:1
A Corynebacterium glutamicum strain with inactivated pyruvate dehydrogenase complex and a deletion of the gene encoding the pyruvate:quinone oxidoreductase
produces about 19 mM l-valine, 28 mM l-alanine and about 55 mM pyruvate from 150 mM glucose. Based on this double mutant C. glutamicum △aceE △pqo, we engineered C. glutamicum for efficient production of pyruvate from glucose by additional deletion of the ldhA gene encoding NAD+-dependent l-lactate dehydrogenase (LdhA) and introduction of a attenuated variant of the acetohydroxyacid synthase (△C–T IlvN). The latter
modification abolished overflow metabolism towards l-valine and shifted the product spectrum to pyruvate production. In shake flasks, the resulting strain C. glutamicum △aceE △pqo △ldhA △C–T ilvN produced about 190 mM pyruvate with a Y
P/S of 1.36 mol per mol of glucose; however, it still secreted significant amounts of l-alanine. Additional deletion of genes encoding the transaminases AlaT and AvtA reduced l-alanine formation by about 50%. In fed-batch fermentations at high cell densities with adjusted oxygen supply during growth
and production (0–5% dissolved oxygen), the newly constructed strain C. glutamicum △aceE △pqo △ldhA △C–T ilvN △alaT △avtA produced more than 500 mM pyruvate with a maximum yield of 0.97 mol per mole of glucose and a productivity of 0.92 mmol g(CDW)−1 h−1 (i.e., 0.08 g g(CDW)
−1 h−1) in the production phase. 相似文献
19.
Simulation and experiment have been used to establish that significant artifacts can be generated in X-pulse CPMG relaxation dispersion experiments recorded on heteronuclear ABX spin-systems, such as 13C
i
–13C
j
–1H, where 13C
i
and 13C
j
are strongly coupled. A qualitative explanation of the origin of these artifacts is presented along with a simple method
to significantly reduce them. An application to the measurement of 1H CPMG relaxation dispersion profiles in an HIV-2 TAR RNA molecule where all ribose sugars are protonated at the 2′ position,
deuterated at all other sugar positions and 13C labeled at all sugar carbons is presented to illustrate the problems that strong 13C–13C coupling introduces and a simple solution is proposed. 相似文献
20.
This study reports the sequence specific chemical shifts assignments for 76 residues of the 94 residues containing monomeric unit of the photosynthetic light-harvesting 2 transmembrane protein complex from Rhodopseudomonas acidophila strain 10050, using Magic Angle Spinning (MAS) NMR in combination with extensive and selective biosynthetic isotope labeling methods. The sequence specific chemical shifts assignment is an essential step for structure determination by MAS NMR. Assignments have been performed on the basis of 2-dimensional proton-driven spin diffusion 13C–13C correlation experiments with mixing times of 20 and 500 ms and band selective 13C–15N correlation spectroscopy on a series of site-specific biosynthetically labeled samples. The decreased line width and the reduced number of correlation signals of the selectively labeled samples with respect to the uniformly labeled samples enable to resolve the narrowly distributed correlation signals of the backbone carbons and nitrogens involved in the long -helical transmembrane segments. Inter-space correlations between nearby residues and between residues and the labeled BChl a cofactors, provided by the 13C–13C correlation experiments using a 500 ms spin diffusion period, are used to arrive at sequence specific chemical shift assignments for many residues in the protein complex. In this way it is demonstrated that MAS NMR methods combined with site-specific biosynthetic isotope labeling can be used for sequence specific assignment of the NMR response of transmembrane proteins. 相似文献