Journal of Biomolecular NMR最新文献_第6页

Band-selective universal 90° and 180° rotation pulses covering the aliphatic carbon chemical shift range for triple resonance experiments on 1.2 GHz spectrometers 用于1.2 GHz光谱仪三重共振实验的90°和180°波段选择性通用旋转脉冲覆盖脂肪碳化学位移范围

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-11-24 DOI: 10.1007/s10858-022-00404-1

Stella Slad, Wolfgang Bermel, Rainer Kümmerle, Daniel Mathieu, Burkhard Luy

{"title":"Band-selective universal 90° and 180° rotation pulses covering the aliphatic carbon chemical shift range for triple resonance experiments on 1.2 GHz spectrometers","authors":"Stella Slad, Wolfgang Bermel, Rainer Kümmerle, Daniel Mathieu, Burkhard Luy","doi":"10.1007/s10858-022-00404-1","DOIUrl":"10.1007/s10858-022-00404-1","url":null,"abstract":"<div>Biomolecular NMR spectroscopy requires large magnetic field strengths for high spectral resolution. Today’s highest fields comprise proton Larmor frequencies of 1.2 GHz and even larger field strengths are to be expected in the future. In protein triple resonance experiments, various carbon bandwidths need to be excited by selective pulses including the large aliphatic chemical shift range. When the spectrometer field strength is increased, the length of these pulses has to be decreased by the same factor, resulting in higher rf-amplitudes being necessary in order to cover the required frequency region. Currently available band-selective pulses like Q3/Q5 excite a narrow bandwidth compared to the necessary rf-amplitude. Because the maximum rf-power allowed in probeheads is limited, none of the selective universal rotation pulses reported so far is able to cover the full (^{13})C aliphatic region on 1.2 GHz spectrometers. In this work, we present band-selective 90° and 180° universal rotation pulses (SURBOP90 and SURBOP180) that have a higher ratio of selective bandwidth to maximum rf-amplitude than standard pulses. Simulations show that these pulses perform better than standard pulses, e. g. Q3/Q5, especially when rf-inhomogeneity is taken into account. The theoretical and experimental performance is demonstrated in offset profiles and by implementing the SURBOP pulses in an HNCACB experiment at 1.2 GHz.</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 5-6","pages":"185 - 195"},"PeriodicalIF":2.7,"publicationDate":"2022-11-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00404-1.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4946477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 2

Pure shift amide detection in conventional and TROSY-type experiments of 13C,15N-labeled proteins 13C, 15n标记蛋白的常规和trosy型实验的纯移位酰胺检测

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-11-18 DOI: 10.1007/s10858-022-00406-z

Jens D. Haller, Andrea Bodor, Burkhard Luy

{"title":"Pure shift amide detection in conventional and TROSY-type experiments of 13C,15N-labeled proteins","authors":"Jens D. Haller, Andrea Bodor, Burkhard Luy","doi":"10.1007/s10858-022-00406-z","DOIUrl":"10.1007/s10858-022-00406-z","url":null,"abstract":"<div>Large coupling networks in uniformly 13C,15N-labeled biomolecules induce broad multiplets that even in flexible proteins are frequently not recognized as such. The reason is that given multiplets typically consist of a large number of individual resonances that result in a single broad line, in which individual components are no longer resolved. We here introduce a real-time pure shift acquisition scheme for the detection of amide protons which is based on 13C-BIRDr,X. As a result the full homo- and heteronuclear coupling network can be suppressed at low power leading to real singlets at substantially improved resolution and uncompromised sensitivity. The method is tested on a small globular and an intrinsically disordered protein (IDP) where the average spectral resolution is increased by a factor of ~ 2 and higher. Equally important, the approach works without saturation of water magnetization for solvent suppression and exchanging amide protons are not affected by saturation transfer.\u0000</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 5-6","pages":"213 - 221"},"PeriodicalIF":2.7,"publicationDate":"2022-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00406-z.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4732483","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Optimizing frequency sampling in CEST experiments 优化CEST实验中的频率采样

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-10-04 DOI: 10.1007/s10858-022-00403-2

Nicolas Bolik-Coulon, D. Flemming Hansen, Lewis E. Kay

{"title":"Optimizing frequency sampling in CEST experiments","authors":"Nicolas Bolik-Coulon, D. Flemming Hansen, Lewis E. Kay","doi":"10.1007/s10858-022-00403-2","DOIUrl":"10.1007/s10858-022-00403-2","url":null,"abstract":"<div>For the past decade chemical exchange saturation transfer (CEST) experiments have been successfully applied to study exchange processes in biomolecules involving sparsely populated, transiently formed conformers. Initial implementations focused on extensive sampling of the CEST frequency domain, requiring significant measurement times. Here we show that the lengthy sampling schemes often used are not optimal and that reduced frequency sampling schedules can be developed without a priori knowledge of the exchange parameters, that only depend on the chosen B1 field, and, to a lesser extent, on the intrinsic transverse relaxation rates of ground state spins. The reduced sampling approach described here can be used synergistically with other methods for reducing measurement times such as those that excite multiple frequencies in the CEST dimension simultaneously, or make use of non-uniform sampling of indirectly detected time domains, to further decrease measurement times. The proposed approach is validated by analysis of simulated and experimental datasets.\u0000</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 5-6","pages":"167 - 183"},"PeriodicalIF":2.7,"publicationDate":"2022-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00403-2.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4186969","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 2

Characterization of conformational heterogeneity via higher-dimensionality, proton-detected solid-state NMR 通过高维质子探测固态核磁共振表征构象非均质性

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-09-23 DOI: 10.1007/s10858-022-00405-0

Ekaterina Burakova, Suresh K. Vasa, Rasmus Linser

{"title":"Characterization of conformational heterogeneity via higher-dimensionality, proton-detected solid-state NMR","authors":"Ekaterina Burakova, Suresh K. Vasa, Rasmus Linser","doi":"10.1007/s10858-022-00405-0","DOIUrl":"10.1007/s10858-022-00405-0","url":null,"abstract":"<div>Site-specific heterogeneity of solid protein samples can be exploited as valuable information to answer biological questions ranging from thermodynamic properties determining fibril formation to protein folding and conformational stability upon stress. In particular, for proteins of increasing molecular weight, however, site-resolved assessment without residue-specific labeling is challenging using established methodology, which tends to rely on carbon-detected 2D correlations. Here we develop purely chemical-shift-based approaches for assessment of relative conformational heterogeneity that allows identification of each residue via four chemical-shift dimensions. High dimensionality diminishes the probability of peak overlap in the presence of multiple, heterogeneously broadened resonances. Utilizing backbone dihedral-angle reconstruction from individual contributions to the peak shape either via suitably adapted prediction routines or direct association with a relational database, the methods may in future studies afford assessment of site-specific heterogeneity of proteins without site-specific labeling.</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 5-6","pages":"197 - 212"},"PeriodicalIF":2.7,"publicationDate":"2022-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00405-0.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4913103","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Correction to: Validation of protein backbone structures calculated from NMR angular restraints using Rosetta 更正:使用Rosetta从核磁共振角度约束计算的蛋白质主结构的验证

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-08-27 DOI: 10.1007/s10858-022-00398-w

Joel Lapin, Alexander A. Nevzorov

引用次数: 0

Measurement of 1Hα transverse relaxation rates in proteins: application to solvent PREs 蛋白质中1Hα横向弛豫速率的测量:在溶剂PREs中的应用

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-08-26 DOI: 10.1007/s10858-022-00401-4

Yuki Toyama, Atul Kaushik Rangadurai, Lewis E. Kay

{"title":"Measurement of 1Hα transverse relaxation rates in proteins: application to solvent PREs","authors":"Yuki Toyama, Atul Kaushik Rangadurai, Lewis E. Kay","doi":"10.1007/s10858-022-00401-4","DOIUrl":"10.1007/s10858-022-00401-4","url":null,"abstract":"<div>It has recently been demonstrated that accurate near surface electrostatic potentials can be calculated for proteins from solvent paramagnetic relaxation enhancements (PREs) of amide protons measured using spin labels of similar structures but different charges (Yu et al. in Proc Natl Acad Sci 118(25):e2104020118, 2021). Here we develop methodology for extending such measurements to intrinsically disordered proteins at neutral pH where amide spectra are of very poor quality. Under these conditions it is shown that accurate PRE values can be measured using the haCONHA experiment that has been modified for recording 1Hα transverse relaxation rates. The optimal pulse scheme includes a spin-lock relaxation element for suppression of homonuclear scalar coupled evolution for all 1Hα protons, except those derived from Ser and Thr residues, and minimizes the radiation damping field from water magnetization that would otherwise increase measured relaxation rates. The robustness of the experiment is verified by developing a second approach using a band selective adiabatic decoupling scheme for suppression of scalar coupling modulations during 1Hα relaxation and showing that the measured PRE values from the two methods are in excellent agreement. The near surface electrostatic potential of a 103-residue construct comprising the C-terminal intrinsically disordered region of the RNA-binding protein CAPRIN1 is obtained at pH 5.5 using both 1HN and 1Hα-based relaxation rates, and at pH 7.4 where only 1Hα rates can be quantified, with very good agreement between potentials obtained under all experimental conditions.\u0000</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 4","pages":"137 - 152"},"PeriodicalIF":2.7,"publicationDate":"2022-08-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"5392647","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 7

The measurement of binding affinities by NMR chemical shift perturbation 用核磁共振化学位移微扰测量结合亲和

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-08-03 DOI: 10.1007/s10858-022-00402-3

Billy Hobbs, Jack Drant, Mike P. Williamson

引用次数: 5

Proton TOCSY NMR relaxation rates quantitate protein side chain mobility in the Pin1 WW domain 质子TOCSY核磁共振弛豫率定量蛋白侧链迁移在Pin1 WW结构域

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-07-21 DOI: 10.1007/s10858-022-00400-5

Gaddafi I. Danmaliki, Peter M. Hwang

{"title":"Proton TOCSY NMR relaxation rates quantitate protein side chain mobility in the Pin1 WW domain","authors":"Gaddafi I. Danmaliki, Peter M. Hwang","doi":"10.1007/s10858-022-00400-5","DOIUrl":"10.1007/s10858-022-00400-5","url":null,"abstract":"<div>Protein side chain dynamics play a vital role in many biological processes, but differentiating mobile from rigid side chains remains a technical challenge in structural biology. Solution NMR spectroscopy is ideally suited for this but suffers from limited signal-to-noise, signal overlap, and a need for fractional 13C or 2H labeling. Here we introduce a simple strategy measuring initial 1H relaxation rates during a 1H TOCSY sequence like DIPSI-2, which can be appended to the beginning of any multi-dimensional NMR sequence that begins on 1H. The TOCSY RF field compels all 1H atoms to behave similarly under the influence of strong coupling and rotating frame cross-relaxation, so that differences in relaxation rates are due primarily to side chain mobility. We apply the scheme to a thermostable mutant Pin1 WW domain and demonstrate that the observed 1H relaxation rates correlate well with two independent NMR measures of side-chain dynamics, cross-correlated 13C relaxation rates in 13CβH2 methylene groups and maximum observable 3J couplings sensitive to the χ1 side chain dihedral angle (3JHα,Hβ, 3JN,Hβ, and 3JCO,Hβ). The most restricted side chains belong to Trp26 and Asn40, which are closely packed to constitute the folding center of the WW domain. None of the other conserved aromatic residues is as immobile as the first tryptophan side chain of the WW domain. The proposed 1H relaxation methodology should make it relatively easy to measure side chain dynamics on uniformly 15N- or 13C-labeled proteins, so long as chemical shift assignments are obtainable.</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 4","pages":"121 - 135"},"PeriodicalIF":2.7,"publicationDate":"2022-07-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00400-5.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4824292","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Distinct stereospecific effect of chiral tether between a tag and protein on the rigidity of paramagnetic tag 标签与蛋白质之间的手性系链对顺磁标签刚性的立体特异性影响

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-07-16 DOI: 10.1007/s10858-022-00399-9

Jia-Liang Chen, Bin Li, Bo Ma, Xun-Cheng Su

{"title":"Distinct stereospecific effect of chiral tether between a tag and protein on the rigidity of paramagnetic tag","authors":"Jia-Liang Chen, Bin Li, Bo Ma, Xun-Cheng Su","doi":"10.1007/s10858-022-00399-9","DOIUrl":"10.1007/s10858-022-00399-9","url":null,"abstract":"<div>Flexibility between the paramagnetic tag and its protein conjugates is a common yet unresolved issue in the applications of paramagnetic NMR spectroscopy in biological systems. The flexibility greatly attenuates the magnetic anisotropy and compromises paramagnetic effects especially for pseudocontact shift and residual dipolar couplings. Great efforts have been made to improve the rigidity of paramagnetic tag in the protein conjugates, however, the effect of local environment vicinal to the protein ligation site on the paramagnetic effects remains poorly understood. In the present work, the stereospecific effect of chiral tether between the protein and a tag on the paramagnetic effects produced by the tag attached via a D- and L-type linker between the protein and paramagnetic metal chelating moiety was assessed. The remarkable chiral effect of the D- and L-type tether between the tag and the protein on the rigidity of paramagnetic tag is disclosed in a number of protein-tag-Ln complexes. The chiral tether formed between the D-type tag and L-type protein surface minimizes the effect of the local environment surrounding the ligation site on the averaging of paramagnetic tag, which is helpful to preserve the rigidity of a paramagnetic tag in the protein conjugates.</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 4","pages":"107 - 119"},"PeriodicalIF":2.7,"publicationDate":"2022-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4939669","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Metabolic15N labeling of the N-glycosylated immunoglobulin G1 Fc with an engineered Saccharomyces cerevisiae strain 用工程酿酒酵母菌株对n -糖基化免疫球蛋白G1 Fc进行代谢标记

IF 2.7 3区生物学

Journal of Biomolecular NMR Pub Date : 2022-07-08 DOI: 10.1007/s10858-022-00397-x

Anjali Shenoy, Alexander R. Davis, Elijah T. Roberts, I. Jonathan Amster, Adam W. Barb

{"title":"Metabolic15N labeling of the N-glycosylated immunoglobulin G1 Fc with an engineered Saccharomyces cerevisiae strain","authors":"Anjali Shenoy, Alexander R. Davis, Elijah T. Roberts, I. Jonathan Amster, Adam W. Barb","doi":"10.1007/s10858-022-00397-x","DOIUrl":"10.1007/s10858-022-00397-x","url":null,"abstract":"<div>The predominant protein expression host for NMR spectroscopy is Escherichia coli, however, it does not synthesize appropriate post-translation modifications required for mammalian protein function and is not ideal for expressing naturally secreted proteins that occupy an oxidative environment. Mammalian expression platforms can address these limitations; however, these are not amenable to cost-effective uniform 15 N labeling resulting from highly complex growth media requirements. Yeast expression platforms combine the simplicity of bacterial expression with the capabilities of mammalian platforms, however yeasts require optimization prior to isotope labeling. Yeast expression will benefit from methods to boost protein expression levels and developing labeling conditions to facilitate growth and high isotope incorporation within the target protein. In this work, we describe a novel platform based on the yeast Saccharomyces cerevisiae that simultaneously expresses the Kar2p chaperone and protein disulfide isomerase in the ER to facilitate the expression of secreted proteins. Furthermore, we developed a growth medium for uniform 15 N labeling. We recovered 2.2 mg/L of uniformly 15 N-labeled human immunoglobulin (Ig)G1 Fc domain with 90.6% 15 N labeling. NMR spectroscopy revealed a high degree of similarity between the yeast and mammalian-expressed IgG1 Fc domains. Furthermore, we were able to map the binding interaction between IgG1 Fc and the Z domain through chemical shift perturbations. This platform represents a novel cost-effective strategy for 15 N-labeled immunoglobulin fragments.</div>","PeriodicalId":613,"journal":{"name":"Journal of Biomolecular NMR","volume":"76 4","pages":"95 - 105"},"PeriodicalIF":2.7,"publicationDate":"2022-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s10858-022-00397-x.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4341619","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0