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1.
Nishit Goradia Amelie Wißbrock Christoph Wiedemann Frank Bordusa Ramadurai Ramachandran Diana Imhof Oliver Ohlenschläger 《Biomolecular NMR assignments》2016,10(2):329-333
Interleukin-36α (IL-36α) is a recently characterised member of the interleukin-1 superfamily. It is involved in the pathogenesis of inflammatory arthritis in one third of psoriasis patients. By binding of IL-36α to its receptor IL-36R via the NF-κB pathway other cytokines involved in inflammatory and apoptotic cascade are activated. The efficacy of complex formation is controlled by N-terminal processing. To obtain a more detailed view on the structure function relationship we performed a heteronuclear multidimensional NMR investigation and here report the 1H, 13C, and 15N resonance assignments for the backbone and side chain nuclei of the pro-inflammatory cytokine interleukin-36α. 相似文献
2.
Meng Zhang Xiao-Wei Yu G. V. T. Swapna Gaohua Liu Rong Xiao Yan Xu Gaetano T. Montelione 《Biomolecular NMR assignments》2018,12(1):63-68
Lipase r27RCL is a 296-residue, 33 kDa monomeric enzyme with high ester hydrolysis activity, which has significant applications in the baking, paper and leather industries. The lipase gene proRCL from Rhizopus microsporus var. chinensis (also Rhizopus chinensis) CCTCC M201021 was cloned as a fusion construct C-terminal to a maltose-binding protein (MBP) tag, and expressed as MBP-proRCL in an Escherichia coli BL21 trxB (DE3) expression system with uniform 2H,13C,15N-enrichment and Ile-δ1, Leu, and Val 13CH3 methyl labeling. The fusion protein was hydrolyzed by Kex2 protease at the recognition site Lys-Arg between residues ?29 and ?28 of the prosequence, producing the enzyme form called r27RCL. Here we report extensive backbone 1H, 15N, and 13C, as well as Ile-δ1, Leu, and Val side chain methyl, NMR resonance assignments for r27RCL. 相似文献
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Koh Takeuchi Dominique P. Frueh Zhen-Yu J. Sun Sebastian Hiller Gerhard Wagner 《Journal of biomolecular NMR》2010,47(1):55-63
We present a 13C direct detection CACA-TOCSY experiment for samples with alternate 13C–12C labeling. It provides inter-residue correlations between 13Cα resonances of residue i and adjacent Cαs at positions i − 1 and i + 1. Furthermore, longer mixing times yield correlations to Cα nuclei separated by more than one residue. The experiment also provides Cα-to-sidechain correlations, some amino acid type identifications and estimates for ψ dihedral angles. The power of the experiment
derives from the alternate 13C–12C labeling with [1,3-13C] glycerol or [2-13C] glycerol, which allows utilizing the small scalar 3JCC couplings that are masked by strong 1JCC couplings in uniformly 13C labeled samples. 相似文献
6.
We present a projected [1H,15N]-HMQC-[1H,1H]-NOESY experiment for observation of NOE interactions between amide protons with degenerate 15N chemical shifts in large molecular systems. The projection is achieved by simultaneous evolution of the multiple quantum
coherence of the nitrogen spin and the attached proton spin. In this way NOE signals can be separated from direct-correlation
peaks also in spectra with low resolution by fully exploiting both 1H and 15N frequency differences, such that sensitivity can be increased by using short maximum evolution times. The sensitivity of
the experiment is not dependent on the projection angle for projections up to 45° and no additional pulses or delays are required
as compared to the conventional 2D [1H,15N]-HMQC-NOESY. The experiment provides two distinct 2D spectra corresponding to the positive and negative angle projections,
respectively. With a linear combination of 1D cross-sections from the two projections the unavoidable sensitivity loss in
projection spectra can be compensated for each particular NOE interaction. We demonstrate the application of the novel projection
experiment for the observation of an NOE interaction between two sequential glycines with degenerate 15N chemical shifts in a 121.3 kDa complex of the linker H1 histone protein with a 152 bp linear DNA. 相似文献
7.
Alejandro Pablo Arena Roxana Piastrellini Gabriela Nuri Barón Bárbara María Civit 《The International Journal of Life Cycle Assessment》2017,22(4):546-556
Purpose
Many tools to quantify the environmental impact of human decisions have been developed, but all of them seem to have a limited application at the regional or local level. A free-of-charge, Argentina-based personal footprint calculator software (YUPI®) has been developed in order to raise awareness among local citizens about the environmental impacts generated by their daily habits. The extensive use of the tool will generate information suitable for future scientific studies based on local data.Methods
The software calculates the ecological, carbon, and water footprints of individuals, implementing specific regional data from Argentina developed by the CLIOPE group, complemented with data from the Water Footprint Network and the Global Footprint Network. The calculator was developed focusing on interface attractiveness, ease of use, language simplicity, and a good trade-off between completion time and fullness.Results and discussion
The YUPI® software allows its users to understand at a glance their contribution to the environmental impacts of modern society and to quantify the reduction opportunities they have at hand. The program’s language and variables reflect local lifestyle choices, making the filling process accessible for children. The calculator was placed online as an educational tool for teachers and students from all educational levels, and it was also used by visitors in local science and educational fairs. Valuable data was collected for future initiatives on impact mitigation.Conclusions
Amplified by the mass media, the new tool has helped raise awareness and discussion about the individual environmental footprint, both in the educational and in the domestic terrain. The strategy of creating a simple, easily administered, and widely available quiz helped bridge the gap between the academy and the people, making available to them the continuously updated information generated by the research groups. This is facilitating citizen not only to understand the complexity of the environmental problems but also to take informed actions leading to their mitigation.8.
Arginine side-chains are often key for enzyme catalysis, protein–ligand and protein–protein interactions. The importance of arginine stems from the ability of the terminal guanidinium group to form many key interactions, such as hydrogen bonds and salt bridges, as well as its perpetual positive charge. We present here an arginine 13Cζ-detected NMR experiment in which a double-quantum coherence involving the two 15Nη nuclei is evolved during the indirect chemical shift evolution period. As the precession frequency of the double-quantum coherence is insensitive to exchange of the two 15Nη; this new approach is shown to eliminate the previously deleterious line broadenings of 15Nη resonances caused by the partially restricted rotation about the Cζ–Nε bond. Consequently, sharp and well-resolved 15Nη resonances can be observed. The utility of the presented method is demonstrated on the L99A mutant of the 19 kDa protein T4 lysozyme, where the measurement of small chemical shift perturbations, such as one-bond deuterium isotope shifts, of the arginine amine 15Nη nuclei becomes possible using the double-quantum experiment. 相似文献
9.
We present two NMR experiments, (3,2)D HNHA and (3,2)D HNHB, for rapid and accurate measurement of 3J(H N-H alpha) and 3J(N-H beta) coupling constants in polypeptides based on the principle of G-matrix Fourier transform NMR spectroscopy and quantitative J-correlation. These experiments, which facilitate fast acquisition of three-dimensional data with high spectral/digital resolution and chemical shift dispersion, will provide renewed opportunities to utilize them for sequence specific resonance assignments, estimation/characterization of secondary structure with/without prior knowledge of resonance assignments, stereospecific assignment of prochiral groups and 3D structure determination, refinement and validation. Taken together, these experiments have a wide range of applications from structural genomics projects to studying structure and folding in polypeptides. 相似文献
10.
Khandekar Jishan Bari Shrikant Sharma Kandala V. R. Chary 《Biomolecular NMR assignments》2018,12(1):51-55
γS-crystallin is a major structural component of the human eye lens, which maintains its stability over the lifetime of an organism with negligible turnover. The G57W mutant of human γS-crystallin (abbreviated hereafter as γS-G57W) is associated with dominant congenital cataracts. In order to provide a structural basis for the ability of γS-G57W causing cataract, we have cloned, overexpressed, isolated and purified the protein. The 2D [15N–1H]-HSQC spectrum recorded with uniformly 13C/15N-labelled γS-G57W was highly dispersed indicating the protein to adopt an ordered conformation. In this paper, we report almost complete sequence-specific 1H, 13C and 15N resonance assignments of γS-G57W using a suite of heteronuclear 3D NMR experiments. 相似文献
11.
MEMBRANE enzymes, because of their lipid content, are insoluble in water and usually solubilized as micelles in aqueous solution by detergents for biochemical study. Direct study of lipid components by means of organic liquids in which they dissolve leads at once to the denaturation of the enzyme. At temperatures appreciably colder than 0° C, however, organic solvents may leave enzymatic activity intact1–3, making it possible to study enzyme reactions. 相似文献
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Roxanne P. Smith Biswaranjan Mohanty Martin L. Williams Martin J. Scanlon Begoña Heras 《Biomolecular NMR assignments》2017,11(2):181-186
DsbD is a disulfide bond reductase present in the inner membrane of many Gamma-Proteobacteria. In the human pathogen Neisseria meningitidis, DsbD is required for viability and represents a potential target for the development of antibiotics. Here we report the chemical shift assignments (HN, N, Cα and Cβ) for the reduced and oxidized forms of the two periplasmic domains of N. meningitidis DsbD, n-NmDsbD and c-NmDsbD. The backbone amide resonances in all four forms were completely assigned, and the secondary structures for the core regions of the proteins were calculated using 13Cαβ shifts. The reduced and oxidized forms of each domain have similar secondary shifts suggesting they retain the same fold. We anticipate that these data will provide an important basis for studying the interaction between n-NmDsbD and c-NmDsbD, which is required for electron transfer across the bacterial cytoplasmic membrane. 相似文献
14.
Muhammad Usman Akbar Tanveer Hussain Bokhari Samina Roohi Khalid Mahmood Zia Mohammad Zuber Nadia Parveen Narmeen Ali 《Russian Journal of Bioorganic Chemistry》2016,42(5):491-496
Human biosynthetic insulin is a polypeptide hormone that plays an important and essential role in control of the level of carbohydrate, protein, and fat metabolism in the blood. Human pancreatic insulin was labeled with 99mTc to form a new radiopharmaceutical with a labeling yield of 99 ± 1% under optimum conditions: 0.1 mL insulin, pH 7, 25 μg stannous chloride, 1 mL (19 mCi) of pertechnetate, room temperature, and 10 min reaction time. The 99mTc–insulin complex was examined using paper chromatography, ITLC, electrophoresis, and HPLC. In addition, in vitro and in vivo study of 99mTc–insulin complex was performed at different time intervals. 相似文献
15.
Jorge A. Vila Pedro Serrano Kurt Wüthrich Harold A. Scheraga 《Journal of biomolecular NMR》2010,48(1):23-30
To evaluate sequential nearest-neighbor effects on quantum-chemical calculations of 13Cα chemical shifts, we selected the structure of the nucleic acid binding (NAB) protein from the SARS coronavirus determined
by NMR in solution (PDB id 2K87). NAB is a 116-residue α/β protein, which contains 9 prolines and has 50% of its residues
located in loops and turns. Overall, the results presented here show that sizeable nearest-neighbor effects are seen only
for residues preceding proline, where Pro introduces an overestimation, on average, of 1.73 ppm in the computed 13Cα chemical shifts. A new ensemble of 20 conformers representing the NMR structure of the NAB, which was calculated with an
input containing backbone torsion angle constraints derived from the theoretical 13Cα chemical shifts as supplementary data to the NOE distance constraints, exhibits very similar topology and comparable agreement
with the NOE constraints as the published NMR structure. However, the two structures differ in the patterns of differences
between observed and computed 13Cα chemical shifts, Δ
ca,i
, for the individual residues along the sequence. This indicates that the Δ
ca,i
-values for the NAB protein are primarily a consequence of the limited sampling by the bundles of 20 conformers used, as
in common practice, to represent the two NMR structures, rather than of local flaws in the structures. 相似文献
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An extension to HN(CO-α/β-N,Cα-J)-TROSY (Permi and Annila in J Biomol NMR 16:221–227, 2000) is proposed that permits the simultaneous determination of the four coupling constants 1
J
N′(i)Cα(i), 2
J
HN(i)Cα(i), 2
J
Cα(i−1)N′(i), and 3
J
Cα(i−1)HN(i) in 15N,13C-labeled proteins. Contrasting the original scheme, in which two separate subspectra exhibit the 2
J
CαN′ coupling as inphase and antiphase splitting (IPAP), we here record four subspectra that exhibit all combinations of inphase
and antiphase splittings possible with respect to both 2
J
CαN′ and 1
J
N′Cα (DIPAP). Complementary sign patterns in the different spectrum constituents overdetermine the coupling constants which can
thus be extracted at higher accuracy than is possible with the original experiment. Fully exploiting data redundance, simultaneous
2D lineshape fitting of the E.COSY multiplet tilts in all four subspectra provides all coupling constants at ultimate precision.
Cross-correlation and differential-relaxation effects were taken into account in the evaluation procedure. By applying a four-point
Fourier transform, the set of spectra is reversibly interconverted between DIPAP and spin-state representations. Methods are
exemplified using proteins of various size. 相似文献
18.
Based on the difference in the CD14 and CD16 expression, two subsets of monocytes were identified in human and other mammalian
blood. These subsets have different patterns of adhesion molecules and chemokine receptors that suggests the different mode
of their interaction with endothelium and tissue traffic. Here, we investigated the ability of CD14+CD16+ and CD14++CD16− monocytes to adhere to endothelial cell monolayer in presence or absence of pro- and anti-inflammatory cytokines. We demonstrated
that CD14+CD16+ monocytes had a higher level of adhesion to intact monolayer of endothelial cells than CD14++CD16− monocytes. Adhesion of CD14++CD16− and CD14+CD16+ monocytes significantly increased in the presence of TNFα or its combination with other cytokines. IFNγ and IL-4 alone did
not affect the adhesion of monocytes. These results show that CD14++CD16− and CD14+CD16+ monocytes can be recruited to the inflamed endothelium, but CD14+CD16+ monocytes adhere to endothelial cells without inflammations twice as strongly as CD14++CD16− monocytes. 相似文献
19.
The 13Cα chemical shifts for 16,299 residues from 213 conformations of four proteins (experimentally determined by X-ray crystallography
and Nuclear Magnetic Resonance methods) were computed by using a combination of approaches that includes, but is not limited
to, the use of density functional theory. Initially, a validation test of this methodology was carried out by a detailed examination
of the correlation between computed and observed 13Cα chemical shifts of 10,564 (of the 16,299) residues from 139 conformations of the human protein ubiquitin. The results of
this validation test on ubiquitin show agreement with conclusions derived from computation of the chemical shifts at the ab initio Hartree–Fock level. Further, application of this methodology to 5,735 residues from 74 conformations of the three remaining
proteins that differ in their number of amino acid residues, sequence and three-dimensional structure, together with a new
scoring function, namely the conformationally averaged root-mean-square-deviation, enables us to: (a) offer a criterion for an accurate assessment of the quality of NMR-derived protein conformations; (b) examine whether X-ray or NMR-solved structures are better representations of the
observed 13Cα chemical shifts in solution; (c) provide evidence indicating that the proposed methodology is more accurate than automated
predictors for validation of protein structures; (d) shed light as to whether the agreement between computed and observed
13Cα chemical shifts is influenced by the identity of an amino acid residue or its location in the sequence; and (e) provide evidence
confirming the presence of dynamics for proteins in solution, and hence showing that an ensemble of conformations is a better
representation of the structure in solution than any single conformation.
Electronic Supplementary Material The online version of this article (doi: ) contains supplementary material, which is available to authorized users. 相似文献
20.
Baraguey C Skouri-Panet F Bontems F Tardieu A Chassaing G Lequin O 《Journal of biomolecular NMR》2004,30(3):385-386