Optimized labeling of 13CHD2 methyl isotopomers in perdeuterated proteins: potential advantages for 13C relaxation studies of methyl dynamics of larger proteins.

Abstract:

:13CHD2 methyl isotopomers are particularly useful to study methyl dynamics in proteins because, as compared with other methyl isotopomers, the 13C relaxation mechanism for this isotopomer is straightforward. However, in the case of proteins, where (omega tau)2 > 1, the refocused INEPT pulse sequence does not completely suppress unwanted 13CH3 signals. The presence of weak 13CH3 peaks is usually not a serious problem for smaller proteins because there are relatively few methyl signals and they are sharp; however, signal overlap becomes more common as the size of the protein increases. We overcome this problem by preparing a protein using a 98% D2O cell culture medium containing 3-(13)C pyruvic acid, 50-60% deuterated at the 3-position, and 4-(13)C 2-ketobutyric acid, 98% and 62% deuterated at the 3- and 4-positions, respectively. This approach significantly reduces the population of the CH3 isotopomer while optimizing the production of 13CHD2, the isotopomer desired for 13C relaxation measurements. In larger proteins where the deuterium T2 may be too short to measure accurately, we also suggest the alternative measurement of the proton T2 of the 13CH2D methyl isotopomer, because these protons are well-isolated from other protons in these highly deuterated samples.

journal_name

J Biomol NMR

authors

Ishima R,Louis JM,Torchia DA

doi

10.1023/a:1012482426306

subject

Has Abstract

pub_date

2001-10-01 00:00:00

pages

167-71

issue

2

eissn

0925-2738

issn

1573-5001

journal_volume

21

pub_type

杂志文章
  • Optimisation of NMR dynamic models I. Minimisation algorithms and their performance within the model-free and Brownian rotational diffusion spaces.

    abstract::The key to obtaining the model-free description of the dynamics of a macromolecule is the optimisation of the model-free and Brownian rotational diffusion parameters using the collected R (1), R (2) and steady-state NOE relaxation data. The problem of optimising the chi-squared value is often assumed to be trivial, ho...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-007-9214-2

    authors: d'Auvergne EJ,Gooley PR

    更新日期:2008-02-01 00:00:00

  • Assignment of cytosine N3 resonances in nucleic acids via intrabase three-bond coupling to amino protons.

    abstract::Coherences were observed between 15N3 of cytosine and its trans amino proton (H42) using a modified gradient-based heteronuclear single quantum coherence (HSQC) pulse sequence optimized for three-bond proton-nitrogen couplings. The method is demonstrated with a 22-nucleotide RNA fragment of the P5abc region of a group...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1008359723515

    authors: Rüdisser S,Pelton JG,Tinoco I Jr

    更新日期:1999-10-01 00:00:00

  • Characterization of the hydrogen bond network in guanosine quartets by internucleotide 3hJ(NC)' and 2hJ(NN) scalar couplings.

    abstract::Scalar coupling correlations across hydrogen bonds with carbonyl groups as acceptors have been observed in a variety of proteins, but not in nucleic acids. Here we present a pulse scheme that allows such an observation and quantification of trans-hydrogen bond 3hJ(NC)' correlations in nucleic acid base pairs, between ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1008307115641

    authors: Dingley AJ,Masse JE,Feigon J,Grzesiek S

    更新日期:2000-04-01 00:00:00

  • Temperature-jump NMR study of protein folding: ribonuclease A at low pH.

    abstract::The kinetic process of folding of bovine pancreatic ribonuclease A in a 2H2O environment at pH 1.2 was examined by a recently developed temperature-jump NMR method (Akasaka et al., (1990) Rev. Sci. Instrum. 61, 66-68). Upon temperature-jump down from 45 degrees C to 29 degrees C, which was attained within 6 s, the pro...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/BF01874569

    authors: Akasaka K,Naito A,Nakatani H

    更新日期:1991-05-01 00:00:00

  • Development and application of aromatic [(13)C, (1)H] SOFAST-HMQC NMR experiment for nucleic acids.

    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

    authors: Sathyamoorthy B,Lee J,Kimsey I,Ganser LR,Al-Hashimi H

    更新日期:2014-11-01 00:00:00

  • 'Random coil' 1H chemical shifts obtained as a function of temperature and trifluoroethanol concentration for the peptide series GGXGG.

    abstract::Proton chemical shifts of a series of disordered linear peptides (H-Gly-Gly-X-Gly-Gly-OH, with X being one of the 20 naturally occurring amino acids) have been obtained using 1D and 2D 1H NMR at pH 5.0 as a function of temperature and solvent composition. The use of 2D methods has allowed some ambiguities in side-chai...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/BF00227466

    authors: Merutka G,Dyson HJ,Wright PE

    更新日期:1995-01-01 00:00:00

  • Selective excitation of intense solvent signals in the presence of radiation damping.

    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

    authors: Otting G,Liepinsh E

    更新日期:1995-06-01 00:00:00

  • Nucleotide-type chemical shift assignment of the encapsulated 40 kbp dsDNA in intact bacteriophage T7 by MAS solid-state NMR.

    abstract::The icosahedral bacteriophage T7 is a 50 MDa double-stranded DNA (dsDNA) virus that infects Escherichia coli. Although there is substantial information on the physical and morphological properties of T7, structural information, based mostly on Raman spectroscopy and cryo-electron microscopy, is limited. Here, we apply...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-014-9840-4

    authors: Abramov G,Goldbourt A

    更新日期:2014-08-01 00:00:00

  • The war of tools: how can NMR spectroscopists detect errors in their structures?

    abstract::Protein structure determination by NMR methods has started in the mid-eighties and has been growing steadily since then. Ca. 14% of the protein structures deposited in the PDB have been solved by NMR. The evaluation of the quality of NMR structures however is still lacking a well-established practice. In this work, we...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-008-9228-4

    authors: Saccenti E,Rosato A

    更新日期:2008-04-01 00:00:00

  • Amino acid type-selective backbone 1H-15N-correlations for Arg and Lys.

    abstract::Four novel amino acid type-selective triple resonance experiments to identify the backbone amino proton and nitrogen resonances of Arg and Lys and of their sequential neighbors in (13C,15N)-labeled proteins are presented: the R(i+1)-HSQC and R(i,i+1)-HSQC select signals originating from Arg side chains, the K(i+1)-HSQ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1011206131623

    authors: Schubert M,Oschkinat H,Schmieder P

    更新日期:2001-08-01 00:00:00

  • Advances towards resonance assignments for uniformly--13C, 15N enriched bacteriorhodopsin at 18.8 T in purple membranes.

    abstract::Solid state NMR spectra from uniformly (13)C, (15)N enriched bacteriorhodospin (bR) purified from H. salinarium were acquired at 18.8 T using magic angle spinning methods. Isolated resonances of 2D (13)C-(13)C spectra exhibited 0.50-0.55 ppm line-widths. Several amino acid types could be assigned, and at least 12 out ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-008-9235-5

    authors: Varga K,Aslimovska L,Watts A

    更新日期:2008-05-01 00:00:00

  • Intensity modulated HSQC and HMQC: two simple methods to measure 3J(HNH)alpha in proteins.

    abstract::Two methods for the measurement of homonuclear 3J(HNH)alpha coupling constants are described. Both HSQC- and HMQC-type experiments employ 'quantitative J-correlation', in which the coupling constant of interest is obtained from the intensity ratio of cross peaks of two spectra. The first spectrum is acquired with 3J(H...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1008343926502

    authors: Permi P,Kilpeläinen I,Annila A,Heikkinen S

    更新日期:2000-01-01 00:00:00

  • GFT projection NMR based resonance assignment of membrane proteins: application to subunit C of E. coli F(1)F (0) ATP synthase in LPPG micelles.

    abstract::G-matrix FT projection NMR spectroscopy was employed for resonance assignment of the 79-residue subunit c of the Escherichia coli F(1)F(0) ATP synthase embedded in micelles formed by lyso palmitoyl phosphatidyl glycerol (LPPG). Five GFT NMR experiments, that is, (3,2)D HNNCO, L-(4,3)D HNNC (alphabeta) C (alpha), L-(4,...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-008-9224-8

    authors: Zhang Q,Atreya HS,Kamen DE,Girvin ME,Szyperski T

    更新日期:2008-03-01 00:00:00

  • Structural investigations of a GYF domain covalently linked to a proline-rich peptide.

    abstract::Protein structure determination of low affinity complexes of interacting macromolecules is often hampered by a lack of observable NOEs between the binding partners. Covalent linkage offers a way to shift the equilibrium of the interaction partners to the bound state. Here we show that a single-chain protein containing...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1024983029700

    authors: Freund C,Kühne R,Park S,Thiemke K,Reinherz EL,Wagner G

    更新日期:2003-10-01 00:00:00

  • Resolving complex mixtures: trilinear diffusion data.

    abstract::Complex mixtures are at the heart of biology, and biomacromolecules almost always exhibit their function in a mixture, e.g., the mode of action for a spider venom is typically dependent on a cocktail of compounds, not just the protein. Information about diseases is encoded in body fluids such as urine and plasma in th...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-013-9752-8

    authors: Björnerås J,Botana A,Morris GA,Nilsson M

    更新日期:2014-04-01 00:00:00

  • Structure and conformation in solution of the parallel-stranded hybrid alpha-d(CGCAATTCGC).beta-d(GCGTTAAGCG) by high-resolution 2D NMR.

    abstract::The unnatural duplex oligonucleotide alpha-d(CGCAATTCGC).beta-d(GCGTTAAGCG) was analyzed by high-resolution NMR methods. All of the exchangeable imino and nonexchangeable protons of the duplex were assigned. Detection of all 10 of the exchangeable imino protons confirms that a parallel, unsymmetrical duplex is formed....

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/BF01875321

    authors: Gmeiner WH,Rayner B,Morvan F,Imbach JL,Lown JW

    更新日期:1992-05-01 00:00:00

  • Selective 1H- 13C NMR spectroscopy of methyl groups in residually protonated samples of large proteins.

    abstract::Methyl (13)CHD(2) isotopomers of all methyl-containing amino-acids can be observed in residually protonated samples of large proteins obtained from [U-(13)C,(1)H]-glucose/D(2)O-based bacterial media, with sensitivity sufficient for a number of NMR applications. Selective detection of some subsets of methyl groups (Ala...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-009-9393-0

    authors: Guo C,Tugarinov V

    更新日期:2010-02-01 00:00:00

  • Optimization of amino acid type-specific 13C and 15N labeling for the backbone assignment of membrane proteins by solution- and solid-state NMR with the UPLABEL algorithm.

    abstract::We present a computational method for finding optimal labeling patterns for the backbone assignment of membrane proteins and other large proteins that cannot be assigned by conventional strategies. Following the approach of Kainosho and Tsuji (Biochemistry 21:6273-6279 (1982)), types of amino acids are labeled with (1...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-010-9462-4

    authors: Hefke F,Bagaria A,Reckel S,Ullrich SJ,Dötsch V,Glaubitz C,Güntert P

    更新日期:2011-02-01 00:00:00

  • Improved 3D gd-HCACO and gd-(H)CACO-TOCSY experiments for isotopically enriched proteins dissolved in H2O.

    abstract::Pulsed field gradients were incorporated into the HCACO experiment for acquiring spectra on isotopically enriched protein samples dissolved in H2O. Excellent water suppression and spectral quality were achieved using the modified pulse sequence (gd-HCACO), as demonstrated for a 13C-/15N-labeled sample of the SH2 domai...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/BF00202041

    authors: Zhang W,Gmeiner WH

    更新日期:1996-05-01 00:00:00

  • Understanding and solving abnormal peak splitting in 3D HCCH-TOCSY and HCC(CO)NH-TOCSY.

    abstract::The 3D HCCH-TOCSY and HCC(CO)NH-TOCSY experiments provide through bond connectivity and are used for side-chain chemical shift assignment by solution-state NMR. Careful design and implementation of the pulse sequence are key to the successful application of the technique particularly when trying to extract the maximum...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-020-00310-4

    authors: Xia Y,Yuwen T,Rossi P

    更新日期:2020-05-01 00:00:00

  • The NMR structure of the domain II of a chloroplastic NifU-like protein OsNifU1A.

    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

    authors: Kumeta H,Ogura K,Asayama M,Katoh S,Katoh E,Teshima K,Inagaki F

    更新日期:2007-06-01 00:00:00

  • Archiving and disseminating integrative structure models.

    abstract::Limitations in the applicability, accuracy, and precision of individual structure characterization methods can sometimes be overcome via an integrative modeling approach that relies on information from all available sources, including all available experimental data and prior models. The open-source Integrative Modeli...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-019-00264-2

    authors: Vallat B,Webb B,Westbrook J,Sali A,Berman HM

    更新日期:2019-07-01 00:00:00

  • Simultaneous detection of amide and methyl correlations using a time shared NMR experiment: application to binding epitope mapping.

    abstract::Simultaneous recording of different NMR parameters is an efficient way to reduce the overall experimental time and speed up structural studies of biological macromolecules. This can especially be beneficial in the case of fast NMR-based drug screening applications or for collecting NOE restraints, where prohibitively ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-007-9178-2

    authors: Würtz P,Aitio O,Hellman M,Permi P

    更新日期:2007-10-01 00:00:00

  • Measuring hydrogen exchange in proteins by selective water saturation in (1)H- (15)N SOFAST/BEST-type experiments: advantages and limitations.

    abstract::HET(ex)-SOFAST NMR (Schanda et al. in J Biomol NMR 33:199-211, 2006) has been proposed some years ago as a fast and sensitive method for semi-quantitative measurement of site-specific amide-water hydrogen exchange effects along the backbone of proteins. Here we extend this concept to BEST readout sequences that provid...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-014-9857-8

    authors: Rennella E,Solyom Z,Brutscher B

    更新日期:2014-11-01 00:00:00

  • Anthranilic acid, the new player in the ensemble of aromatic residue labeling precursor compounds.

    abstract::The application of metabolic precursors for selective stable isotope labeling of aromatic residues in cell-based protein overexpression has already resulted in numerous NMR probes to study the structural and dynamic characteristics of proteins. With anthranilic acid, we present the structurally simplest precursor for ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-017-0129-2

    authors: Schörghuber J,Geist L,Bisaccia M,Weber F,Konrat R,Lichtenecker RJ

    更新日期:2017-09-01 00:00:00

  • Structural features of the metal binding site and dynamics of gallium putidaredoxin, a diamagnetic derivative of a Cys4Fe2S2 ferredoxin.

    abstract::The first reconstitution of an Fe2S2 ferredoxin with a diamagnetic prosthetic group was recently described [Kazanis et al. (1995) J. Am. Chem. Soc., 117, 6625-6626]. The replacement of the iron-sulfur cluster of the bacterial ferredoxin putidaredoxin (Pdx) by gallium (Ga3+) renders the protein diamagnetic and permits ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1023/a:1018369721091

    authors: Kazanis S,Pochapsky TC

    更新日期:1997-06-01 00:00:00

  • Strategy for automated NMR resonance assignment of RNA: application to 48-nucleotide K10.

    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

    authors: Krähenbühl B,Lukavsky P,Wider G

    更新日期:2014-08-01 00:00:00

  • Secondary H/D isotope effect on hydrogen-bonded hydroxyl groups as a tool for recognizing distance constraints in conformational analysis of oligosaccharides.

    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

    authors: Dabrowski J,Grosskurth H,Baust C,Nifant'ev NE

    更新日期:1998-07-01 00:00:00

  • Cell signaling, post-translational protein modifications and NMR spectroscopy.

    abstract::Post-translationally modified proteins make up the majority of the proteome and establish, to a large part, the impressive level of functional diversity in higher, multi-cellular organisms. Most eukaryotic post-translational protein modifications (PTMs) denote reversible, covalent additions of small chemical entities ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-012-9674-x

    authors: Theillet FX,Smet-Nocca C,Liokatis S,Thongwichian R,Kosten J,Yoon MK,Kriwacki RW,Landrieu I,Lippens G,Selenko P

    更新日期:2012-11-01 00:00:00

  • A simple protocol for expression of isotope-labeled proteins in Escherichia coli grown in shaker flasks at high cell density.

    abstract::Protein expression in E. coli grown in shaker flasks is a routine and pivotal tool in many research laboratories. To maximize protein yields, cells are normally induced in the middle of the linear growth phase, typically at an OD600 of ≤ 1 for cells grown in Luria-Bertani (LB) medium at 37 °C. We recently showed that ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章

    doi:10.1007/s10858-019-00285-x

    authors: Cai M,Huang Y,Craigie R,Clore GM

    更新日期:2019-12-01 00:00:00