Abstract:
:Silkworms serve as promising bioreactors for the production of recombinant proteins, including glycoproteins and membrane proteins, for structural and functional protein analyses. However, lack of methodology for stable isotope labeling has been a major deterrent to using this expression system for nuclear magnetic resonance (NMR) structural biology. Here we developed a metabolic isotope labeling technique using commercially available silkworm larvae. The fifth instar larvae were infected with baculoviruses for co-expression of recombinant human immunoglobulin G (IgG) as a test molecule, with calnexin as a chaperone. They were subsequently reared on an artificial diet containing (15)N-labeled yeast crude protein extract. We harvested 0.1 mg of IgG from larva with a (15)N-enrichment ratio of approximately 80%. This allowed us to compare NMR spectral data of the Fc fragment cleaved from the silkworm-produced IgG with those of an authentic Fc glycoprotein derived from mammalian cells. Therefore, we successfully demonstrated that our method enables production of isotopically labeled glycoproteins for NMR studies.
journal_name
J Biomol NMRjournal_title
Journal of biomolecular NMRauthors
Yagi H,Nakamura M,Yokoyama J,Zhang Y,Yamaguchi T,Kondo S,Kobayashi J,Kato T,Park EY,Nakazawa S,Hashii N,Kawasaki N,Kato Kdoi
10.1007/s10858-015-9930-ysubject
Has Abstractpub_date
2015-06-01 00:00:00pages
157-67issue
2eissn
0925-2738issn
1573-5001journal_volume
62pub_type
杂志文章abstract::Selective water excitation schemes are provided which rely on the radiation damping effect in probeheads characterized by high quality factors. The schemes are implemented in homonuclear NOE and ROE experiments, designed for the selective observation of water-protein cross peaks and their assignment using standard pro...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/BF00182286
更新日期:1995-06-01 00:00:00
abstract::An 'isotopomer-selected NOE' (ISNOE) method for the unequivocal identification of mutually hydrogen-bond-linked hydroxyl groups is described. It relies on the fact that the OH group's signal patterns obtained for a partially deuterated sample originate from both isotopomers of the 'partner' hydroxyl, whereas a NOE for...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/A:1008237308320
更新日期:1998-07-01 00:00:00
abstract::We discuss the optimum experimental conditions to obtain assignment spectra for solid proteins at magic-angle spinning (MAS) frequencies around 100 kHz. We present a systematic examination of the MAS dependence of the amide proton T 2' times and a site-specific comparison of T 2' at 93 kHz versus 60 kHz MAS frequency....
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-015-9975-y
更新日期:2015-10-01 00:00:00
abstract::Staphylococcus aureus hibernation promoting factor (SaHPF) is a 22,2 kDa protein which plays a crucial role in 100S Staphylococcus aureus ribosome formation during stress. SaHPF consists of N-terminal domain (NTD) that prevents proteins synthesis by binding to the 30S subunit at the P- and A-sites, connected through a...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-019-00254-4
更新日期:2019-05-01 00:00:00
abstract::The arrival of very high field magnets and cryogenic circuitries, and the development of relaxation-optimized pulse sequences have added powerful tools for increasing sensitivity and resolution in NMR studies of biomacromolecules. The potential of these advances is not fully realized in practice, however, since curren...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章,评审
doi:10.1023/B:JNMR.0000042946.04002.19
更新日期:2004-09-01 00:00:00
abstract::As part of efforts to develop improved methods for NMR protein sample preparation and structure determination, the Northeast Structural Genomics Consortium (NESG) has implemented an NMR screening pipeline for protein target selection, construct optimization, and buffer optimization, incorporating efficient microscale ...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-009-9386-z
更新日期:2010-01-01 00:00:00
abstract::The use of dipolar shifts as important constraints in refining molecular structure of paramagnetic metalloproteins by solution NMR is now well established. A crucial initial step in this procedure is the determination of the orientation. of the anisotropic paramagnetic susceptibility tensor in the molecular frame whic...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/a:1008309121949
更新日期:2000-06-01 00:00:00
abstract::We have analyzed the relaxation properties of all (31)P nuclei in an RNA cUUCGg tetraloop model hairpin at proton magnetic field strengths of 300, 600 and 900 MHz in solution. Significant H, P dipolar contributions to R (1) and R (2) relaxation are observed in a protonated RNA sample at 600 MHz. These contributions ca...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-009-9343-x
更新日期:2009-09-01 00:00:00
abstract::NifU-like proteins are a highly conserved protein that serves as the scaffold for assembly of Fe-S clusters. Chloroplastic NifU-like proteins have tandem NifU like domains, named domain I and domain II. Although the amino acid sequences of these domains are very similar to each other, the predicted functional region f...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-007-9155-9
更新日期:2007-06-01 00:00:00
abstract::NMR View is a computer program designed for the visualization and analysis of NMR data. It allows the user to interact with a practically unlimited number of 2D, 3D and 4D NMR data files. Any number of spectral windows can be displayed on the screen in any size and location. Automatic peak picking and facilitated peak...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/BF00404272
更新日期:1994-09-01 00:00:00
abstract::Residual dipolar couplings provide complementary information to the nuclear Overhauser effect measurements that are traditionally used in biomolecular structure determination by NMR. In a de novo structure determination, however, lack of knowledge about the degree and orientation of molecular alignment complicates the...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-007-9215-1
更新日期:2008-02-01 00:00:00
abstract::Higher sensitivity of NMR spectrometers and novel isotopic labeling schemes have ushered the development of rapid data acquisition methodologies, improving the time resolution with which NMR data can be acquired. For nucleic acids, longitudinal relaxation optimization in conjunction with Ernst angle excitation (SOFAST...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-014-9856-9
更新日期:2014-11-01 00:00:00
abstract::The ability to simultaneously measure many long-range distances is critical to efficient and accurate determination of protein structures by solid-state NMR (SSNMR). So far, the most common distance constraints for proteins are 13C-15N distances, which are usually measured using the rotational-echo double-resonance (R...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-018-0187-0
更新日期:2018-05-01 00:00:00
abstract::A large number of amino acids other than the canonical amino acids can now be easily incorporated in vivo into proteins at genetically encoded positions. The technology requires an orthogonal tRNA/aminoacyl-tRNA synthetase pair specific for the unnatural amino acid that is added to the media while a TAG amber or frame...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章,评审
doi:10.1007/s10858-009-9365-4
更新日期:2010-01-01 00:00:00
abstract::A procedure is presented for automated sequence-specific assignment of NMR resonances of uniformly [(13)C, (15)N]-labeled RNA. The method is based on a suite of four through-bond and two through-space high-dimensional automated projection spectroscopy (APSY) experiments. The approach is exemplified with a 0.3 mM sampl...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-014-9841-3
更新日期:2014-08-01 00:00:00
abstract::Understanding of the molecular mechanisms of protein function requires detailed insight into the conformational landscape accessible to the protein. Conformational changes can be crucial for biological processes, such as ligand binding, protein folding, and catalysis. NMR spectroscopy is exquisitely sensitive to such ...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-018-0193-2
更新日期:2018-06-01 00:00:00
abstract::New base-type-edited transverse-relaxation optimized CT-HCN(C) experiments are presented that yield intra-base and sugar-to-base correlations for 13C-15N labeled RNA. High spectral resolution in the 13C and 15N dimensions is achieved by constant time (CT) frequency editing. A spectral editing filter applied during the...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-005-8872-1
更新日期:2005-08-01 00:00:00
abstract::We perform a detailed comparison of fast backbone dynamics probed at amide nitrogen versus carbonyl carbon sites for dematin headpiece C-terminal domain (DHP) and its S74E mutant (DHPS74E). Carbonyl dynamics is probed via auto-correlated longitudinal rates and transverse C'/C'-C(alpha) CSA/dipolar and C'/C'-N CSA/dipo...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-010-9417-9
更新日期:2010-06-01 00:00:00
abstract::Protein-ligand titrations can readily be monitored with a trimethylsilyl (TMS) tag. Owing to the intensity, narrow line shape and unique chemical shift of a TMS group, dissociation constants can be determined from straightforward 1D 1H-NMR spectra not only in the fast but also in the slow exchange limit. The tag is ea...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-018-0173-6
更新日期:2018-04-01 00:00:00
abstract::The biological environment in which a protein performs its function is a crowded milieu containing millions of molecules that can potentially lead to a great many transient, non-specific interactions. NMR spectroscopy is especially well suited to study these weak molecular contacts. Here, non-specific interactions bet...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-013-9705-2
更新日期:2013-03-01 00:00:00
abstract::Under the condition that the longitudinal relaxation time of spin I is shorter than the longitudinal relaxation time of spin S the steady-state magnetization in [S,I]-TROSY-type experiments can be enhanced by intermediate storage of a part of the steady-state magnetization of spin I on spin S with a pulse sequence ele...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/a:1012431013082
更新日期:2001-10-01 00:00:00
abstract::This paper describes the use of single transition-to-single transition polarization transfer (ST2-PT) in transverse relaxation-optimized spectroscopy (TROSY), where it affords a [Formula: see text] sensitivity enhancement for kinetically stable amide 15N-1H groups in proteins. Additional, conventional improvements of ...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/A:1008268930690
更新日期:1998-08-01 00:00:00
abstract::Fast multidimensional NMR with a time resolution of a few seconds provides a new tool for high throughput screening and site-resolved real-time studies of kinetic molecular processes by NMR. Recently we have demonstrated the feasibility to record protein 1H-15N correlation spectra in a few seconds of acquisition time ...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-005-4425-x
更新日期:2005-12-01 00:00:00
abstract::Detection of (15)N in multidimensional NMR experiments of proteins has sparsely been utilized because of the low gyromagnetic ratio (γ) of nitrogen and the presumed low sensitivity of such experiments. Here we show that selecting the TROSY components of proton-attached (15)N nuclei (TROSY (15)NH) yields high quality s...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-015-9991-y
更新日期:2015-12-01 00:00:00
abstract::A new method is proposed for docking ligands into proteins in cases where an NMR-determined solution structure of a related complex is available. The method uses a set of experimentally determined values for protein-ligand, ligand-ligand, and protein-protein restraints for residues in or near to the binding site, comb...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/a:1008379225053
更新日期:1999-06-01 00:00:00
abstract::A computer program has been developed to accurately and automatically predict the 1H and 13C chemical shifts of unassigned proteins on the basis of sequence homology. The program (called SHIFTY) uses standard sequence alignment techniques to compare the sequence of an unassigned protein against the BioMagResBank--a pu...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1023/a:1018373822088
更新日期:1997-12-01 00:00:00
abstract::No matter the source of compounds, drug discovery campaigns focused directly on the target are entirely dependent on a consistent stream of reliable data that reports on how a putative ligand interacts with the protein of interest. The data will derive from many sources including enzyme assays and many types of biophy...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-020-00343-9
更新日期:2020-11-01 00:00:00
abstract::Simple pulse schemes are presented for the measurement of methyl (13)C and (1)H CSA values from (1)H-(13)C dipole/(13)C CSA and (1)H-(13)C dipole/(1)H CSA cross-correlated relaxation. The methodology is applied to protein L and malate synthase G. Average (13)C CSA values are considerably smaller for Ile than Leu/Val (...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-004-4349-x
更新日期:2004-12-01 00:00:00
abstract::Signal overlapping is a major bottleneck for protein NMR analysis. We propose a new method, stable-isotope-assisted parameter extraction (SiPex), to resolve overlapping signals by a combination of amino-acid selective isotope labeling (AASIL) and tensor decomposition. The basic idea of Sipex is that overlapping signal...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-019-00295-9
更新日期:2020-03-01 00:00:00
abstract::The NMR derived translational diffusion coefficients were performed on unlabeled and uniformly labeled 13C,15N human insulin in water, both in neat, with zinc ions only, and in pharmaceutical formulation, containing only m-cresol as phenolic ligand, glycerol and zinc ions. The results show the dominant role of the pH ...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/s10858-018-0197-y
更新日期:2018-06-01 00:00:00