Concentrated Solution Effects

Warren, Warren S.

doi:10.1002/9780470034590.emrstm0088.pub2

Cited by 5 publications

(8 citation statements)

References 49 publications

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“…Expression and puri cation of ubiquitin K48C mutant. Uniformly 15 N labeled K48C ubiquitin mutant (Ubi K48C ) was expressed and puri ed as reported by Olivier et al 32 . Brie y, the recombinant human K48C mutant was generated from the ubiquitin wild-type gene (pRSET) vector using the QuickChange® kit from Stratagene (CA, USA).…”

Section: Methodsmentioning

confidence: 99%

“…Brie y, the recombinant human K48C mutant was generated from the ubiquitin wild-type gene (pRSET) vector using the QuickChange® kit from Stratagene (CA, USA). The protein was expressed in E. coli BL21 (DE3) cells cultured at 30°C in M9 minimal media, containing 15 NH 4 Cl (Cambridge Isotope Laboratories Inc.) salt as the only nitrogen source. Protein overexpression was induced at an optical density (OD 600 ) above 0.8 by adding 1 mM of isopropyl β-D-1-thiogalactopyranoside (IPTG) and was carried out for 5 hours.…”

Section: Methodsmentioning

confidence: 99%

“…The puri ed protein was concentrated, lyophilized, and stored at -20°C. For NMR samples, 15 N Ubi K48C was solubilized in 10 mM sodium acetate buffer (pH 6.0), 100 mM NaN 3 and 2 mM dithiothreitol (DTT) with to nal protein concentration of 0.5 mM.…”

Section: Methodsmentioning

confidence: 99%

“…Recombinant Raf kinase inhibitor protein (RKIP) was expressed and puri ed as reported previously by Lee et al 33 . Transformed E. coli BL21(DE3) pLysS cells (Invitrogen™) were grown in M9 minimal media containing 15 NH 4 Cl (Cambridge Isotope Laboratories Inc.), at 30°C. Protein overexpression was initiated by adding 0.4 mM IPTG when the OD 600 was above 1.1 and was carried out for 5 hours.…”

Section: Methodsmentioning

confidence: 99%

“…Some of these pulse schemes also reduce the unwanted exchange between labile sites (e.g., amide protons) and water, increasing the intensity of NOE cross peaks. In addition, high and ultra-high magnetic elds enhance the longitudinal relaxation of the spin magnetization and augment NOE sensitivity; yet, radiation damping complicates water suppression 14,15 .…”

Section: Introductionmentioning

confidence: 99%

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Water irradiation devoid pulses enhance the sensitivity of 1H,1H nuclear Overhauser effects

Manu

Olivieri²,

Veglia³

2022

J Biomol NMR

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The nuclear Overhauser effect (NOE) is one of NMR spectroscopy's most important and versatile parameters. NOE is routinely utilized to determine the structures of medium-to-large size biomolecules and characterize protein-protein, protein-RNA, protein-DNA, and protein-ligand interactions in aqueous solutions. Typical [1H,1H] NOESY pulse sequences incorporate water suppression schemes to reduce the water signal that dominates 1H-detected spectra and minimize NOE intensity losses due to unwanted polarization exchange between water and labile protons. However, at high-and ultra-high magnetic elds, the excitation of the water signal during the execution of the NOESY pulse sequences may cause signi cant attenuation of NOE cross-peak intensities. Using an evolutionary algorithm coupled with arti cial intelligence, we recently designed high-delity pulses [Water irrAdiation DEvoid (WADE) pulses] that elude water excitation and irradiate broader bandwidths relative to commonly used pulses. Here, we demonstrate that WADE pulses, implemented into the 2D [1H,1H] NOESY experiments, increase the intensity of the NOE cross-peaks for labile and, to a lesser extent, non-exchangeable protons. We applied the new 2D [1H,1H] WADE-NOESY pulse sequence to two well-folded, medium-size proteins, i.e., the K48C mutant of ubiquitin and the Raf kinase inhibitor protein (RKIP). We observed a net increase of the NOE intensities varying from 30 to 170% compared to the commonly used NOESY experiments. The new WADE pulses can be easily engineered into 2D and 3D homo-and hetero-nuclear NOESY pulse sequences to boost their sensitivity.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Water irradiation devoid pulses enhance the sensitivity of 1H,1H nuclear Overhauser effects

Manu

Olivieri²,

Veglia³

2022

J Biomol NMR

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Detection of iron-labeled single cells by MR imaging based on intermolecular double quantum coherences at 14 T

Cho

Hong

Cho

et al. 2012

Journal of Magnetic Resonance

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To evaluate the efficiency and feasibility of intermolecular multiple quantum coherence (iMQC) magnetic resonance (MR) imaging for single cell detection, we obtained intermolecular double quantum coherence (iDQC) and conventional gradient echo (GE) images of macrophage cells labeled by contrast agents in gel. The iDQC images obtained with echo-planar readout visualized the labeled cells effectively and with a higher contrast than seen in conventional GE images, especially at low planar resolutions and with thick slices. This implies that iDQC imaging with contrast agents could be a good alternative to conventional MR imaging for detecting labeled single cells or cell tracking under favorable conditions.

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Revisiting the mean-field picture of dipolar effects in solution NMR

Chen

Branca

Warren

2012

The Journal of Chemical Physics

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For more than three decades, the classical or mean-field picture describing the distant dipolar field has been almost always simplified to an effective field proportional to the local longitudinal magnetization, differing only by a scale factor of 1.5 for homomolecular (identical resonance frequency) and heteromolecular interactions. We re-examine the underlying assumptions, and show both theoretically and experimentally that the mathematical framework needs to be modified for modern applications such as imaging. We demonstrate new pulse sequences which produce unexpected effects; for example, modulating an arbitrarily small fraction of the magnetization can substantially alter the frequency evolution. Thus, matched gradient pulse pairs (a seemingly innocuous module in thousands of existing pulse sequences) can alter the time evolution in highly unexpected ways, particularly with small flip angle pulses such as those used in hyperpolarized experiments. We also show that specific gradient pulse combinations can retain only dipolar interactions between unlike spins, and the dipolar field can generate a secular Hamiltonian proportional to I x .

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Concentrated Solution Effects

Cited by 5 publications

References 49 publications

Water irradiation devoid pulses enhance the sensitivity of 1H,1H nuclear Overhauser effects

Water irradiation devoid pulses enhance the sensitivity of 1H,1H nuclear Overhauser effects

Detection of iron-labeled single cells by MR imaging based on intermolecular double quantum coherences at 14 T

Revisiting the mean-field picture of dipolar effects in solution NMR

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