Improved signal fidelity in 4-pulse DEER with Gaussian pulses

Teucher, Markus; Bordignon, Enrica

doi:10.1016/j.jmr.2018.09.003

Cited by 49 publications

(45 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Q‐band DEER measurements were performed at 50 K on a Bruker ELEXSYS E580 with a separate electron–electron double resonance (ELDOR) source or an arbitrary waveform generator (AWG) with a 150 W traveling wave tube (TWT) amplifier and a homemade resonator. A 4‐pulse DEER sequence with 100 MHz separation between the pulses was applied with 12 ns rectangular pulses (for T4 lysozyme) or 14 ns full width at half maximum Gaussian pulses (for Bid) for both observer and pump frequencies . Additional artefacts in the DEER trace from the coherent AWG generated pulses were removed by 16‐step phase cycling.…”

Section: Methodsmentioning

confidence: 99%

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

et al. 2019

Self Cite

View full text Add to dashboard Cite

The availability of bioresistant spin labels is crucial for the optimization of site‐directed spin labeling protocols for EPR structural studies of biomolecules in a cellular context. As labeling can affect proteins’ fold and/or function, having the possibility to choose between different spin labels will increase the probability to produce spin‐labeled functional proteins. Here, we report the synthesis and characterization of iodoacetamide‐ and maleimide‐functionalized spin labels based on the gem ‐diethyl pyrroline structure. The two nitroxide labels are compared to conventional gem ‐dimethyl analogs by site‐directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy, using two water soluble proteins: T4 lysozyme and Bid. To foster their use for structural studies, we also present rotamer libraries for these labels, compatible with the MMM software. Finally, we investigate the “true” biocompatibility of the gem ‐diethyl probes comparing the resistance towards chemical reduction of the NO group in ascorbate solutions and E. coli cytosol at different spin concentrations.

show abstract

Section: Methodsmentioning

confidence: 99%

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

et al. 2019

Self Cite

View full text Add to dashboard Cite

show abstract

“…The sidebands of the sinc-shaped excitation profile of 15 rectangular pulses increase the overlap of the observer and pump pulse in DEER, resulting in so called '2+1' artefacts at the end of the DEER trace. It has recently been shown that those artefacts can be reduced by replacing the rectangular pulses with Gaussian pulses (Teucher and Bordignon, 2018). Gaussian pulses also have a frequency function of ( ) = 0 but an amplitude function:…”

Section: Open Accessmentioning

confidence: 99%

“…20 FWHM describes the full width at half maximum of the pulse in the time domain (Teucher and Bordignon, 2018). During a rectangular or Gaussian pulse the magnetisation vector is rotated around the 1 field with an angle that is independent of the initial orientation of the magnetisation vector.…”

Section: Open Accessmentioning

confidence: 99%

“…Increasing the sensitivity of DEER spectroscopy is an active field of research (Borbat et al, 2013;Breitgoff et al, 2017;Doll et al, 2015;Jeschke et al, 2004;Lovett et al, 2012;Milikisiyants et al, 2019;Polyhach et al, 2012;Tait and Stoll, 2016;Teucher and Bordignon, 2018). A very elegant approach to increasing DEER sensitivity has been made possible by the availability of arbitrary waveform generators with time resolution in the nanosecond region as they allow the generation of 30 broadband microwave pulses (Doll et al, 2013;Doll and Jeschke, 2017;Spindler et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

“…A calculation shows what performance of shaped pulses can be expected for different trace lengths and spin concentrations. 35 https://doi.org/10.5194/mr-2020-5…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Optimising broadband pulses for DEER depends on concentration and distance range of interest

Scherer¹,

Tischlik²,

Weickert³

et al. 2020

Preprint

View full text Add to dashboard Cite

Abstract. EPR distance determination in the nanometre region has become an important tool for studying the structure and interaction of macromolecules. Arbitrary waveform generators (AWGs), which have recently become commercially available for EPR spectrometers, have the potential to increase the sensitivity of the most common technique double electron-electron resonance (DEER, also called PELDOR), as they allow the generation of broadband pulses. There are several families of broadband pulses, which are different in general pulse shape and the parameters that define them. Here, we compare the most common broadband pulses. When broadband pulses lead to a larger modulation depth they also increase the background decay of the DEER trace. Depending on the dipolar evolution time this can significantly increase the noise level towards the end of the form factor and limit the potential increase of the modulation to noise ratio (MNR). We found asymmetric hyperbolic secant (HS{1,6}) pulses to perform best for short DEER traces leading to a MNR improvement of up to 86 % compared to rectangular pulses. For longer traces we found symmetric hyperbolic secant (HS{1,1}) pulses to perform best, however, the increase compared to rectangular pulses goes down to 43 %.

show abstract

Protein‐Protein Interaction and Conformational Change in the Alpha‐Helical Membrane Transporter BtuCD‐F in the Native Cellular Envelope

Joseph

2024

ChemBioChem

View full text Add to dashboard Cite

Alpha‐helical membrane proteins perform numerous critical functions essential for the survival of living organisms. Traditionally, these proteins are extracted from membranes using detergent solubilization and reconstitution into liposomes or nanodiscs. However, these processes often obscure the effects of nanoconfinement and the native environment on the structure and conformational heterogeneity of the target protein. We demonstrate that pulsed dipolar electron spin resonance spectroscopy, combined with the Gd³⁺‐nitroxide spin pair, enables the selective observation of the vitamin B12 importer BtuCD‐F in its native cellular envelope. Despite the high levels of non‐specific labeling in the envelope, this orthogonal approach combined with the long phase‐memory time for the Gd³⁺ spin enables the observation of the target protein complex at a few micromolar concentrations with high resolution. In the native envelope, vitamin B12induces a distinct conformational shift at the BtuCD‐BtuF interface, which is not observed in the micelles. This approach offers a general strategy for investigating protein‐protein and protein‐ligand/drug interactions and conformational changes of the alpha‐helical membrane proteins in their native envelope context.

show abstract

Improved signal fidelity in 4-pulse DEER with Gaussian pulses

Cited by 49 publications

References 40 publications

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

Optimising broadband pulses for DEER depends on concentration and distance range of interest

Protein‐Protein Interaction and Conformational Change in the Alpha‐Helical Membrane Transporter BtuCD‐F in the Native Cellular Envelope

Contact Info

Product

Resources

About