A dual-mode microwave resonator for double electron–electron spin resonance spectroscopy at W-band microwave frequencies

Tkach, Igor; Sicoli, Giuseppe; Höbartner, Claudia; Bennati, Marina

doi:10.1016/j.jmr.2011.01.012

Cited by 32 publications

(23 citation statements)

References 22 publications

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“…This problem was overcome with the introduction of Gd pairs as spin labels [36,37], eliminating orientation selection due to the essentially isotropic g factor of Gd 3+ The Gd 3+ can be introduced in the side chain C1 (Figure 1g) [38••] but other side chain chelates are possible [37,39,40•]. Gd 3+ –nitroxide pairs have also been investigated with success [39,40•], particularly when combined with a recently developed bimodal resonator for W band [41] that allows realization of the full potential of the method [38••]. Although orientation selection is reintroduced when the nitroxide is rigid, only two angles are required to describe the geometry and these can be extracted from the data combined with simple modeling.…”

Section: Advances In Distance Determinationmentioning

confidence: 99%

Technological advances in site-directed spin labeling of proteins

Hubbell

López

Altenbach

et al. 2013

Current Opinion in Structural Biology

283

300

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Molecular flexibility over a wide time range is of central importance to the function of many proteins, both soluble and membrane. Revealing the modes of flexibility, their amplitudes, and time scales under physiological conditions is the challenge for spectroscopic methods, one of which is site-directed spin labeling EPR (SDSL-EPR). Here we provide an overview of some recent technological advances in SDSL-EPR related to investigation of structure, structural heterogeneity, and dynamics of proteins. These include new classes of spin labels, advances in measurement of long range distances and distance distributions, methods for identifying backbone and conformational fluctuations, and new strategies for determining the kinetics of protein motion.

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Section: Advances In Distance Determinationmentioning

confidence: 99%

Technological advances in site-directed spin labeling of proteins

Hubbell

López

Altenbach

et al. 2013

Current Opinion in Structural Biology

283

300

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“…16 Due to its large D value, the sensitivity of DEER experiments obtained with the Mn 2+ -EDTA tag is less than what has been obtained with the Gd 3+ -DOTA-amide tag. 16 This situation, however, may change completely if tags with low D values become available, in particular, in combination with a bimodal cavity 28 that allows placement of the pump and observe pulses on two neighboring narrow high-intensity hyperfine components (separated by ∼280 MHz).…”

Section: +mentioning

confidence: 99%

Nanometer-Range Distance Measurement in a Protein Using Mn²⁺ Tags

Banerjee

Yagi

Huber

et al. 2012

J. Phys. Chem. Lett.

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Pulse electron paramagnetic resonance measurements of long-range (nm scale) distances between spin labels site-specifically attached to biomacromolecules have proven highly effective in structural studies. The most commonly used spin labels are stable nitroxide radicals, and measurements are usually carried out at X-band frequencies (∼9.5 GHz, 0.35 T). Higher magnetic fields open new possibilities for distance measurements with increased sensitivity using alternative spin labels containing half-integer high-spin metal ions. Here we demonstrate W-band (95 GHz) pulse double electron−electron resonance (DEER) distance measurements in a protein labeled with two Mn 2+ -EDTA tags. The distance distribution obtained is in excellent agreement with model calculations based on the known solution NMR structure. Thus, sitespecific labeling with Mn 2+ tags opens a highly promising approach to nanometer distance measurements in biological macromolecules. SECTION: Biophysical Chemistry L ong-range (nm scale) distance measurements between specific sites in biological macromolecules offer important insight into their structure and interactions. In the past decade, such distance measurements by pulse electron paramagnetic resonance (EPR) techniques, often referred to as pulse dipolar spectroscopy (PDS), have proven highly efficient for nitroxidelabeled proteins and nucleic acids.1−3 Distances in the range of 2−5 nm can be routinely accessed, and distances up to 8 nm can be determined under favorable conditions. 4,5 In proteins, site-directed spin labeling is usually achieved by covalent attachment of nitroxide compounds to cysteine residues that are either native or, more commonly, have been introduced at strategically chosen positions by site-directed mutagenesis. where r is the electron−electron distance and θ is the angle between the interelectron vector r and the external magnetic field. The measurements are carried out at low temperatures on frozen solutions. The most popular PDS experiment is the four-pulse double electron−electron resonance (DEER) sequence, 7 often also referred to as PELDOR (pulse electron double resonance). DEER carried out at standard X-band frequencies (∼9.5 GHz, 0.35 T) has become well-established. An order of magnitude increase in sensitivity can be gained by high-frequency/highfield DEER, provided that the microwave (mw) power is sufficient to generate short enough microwave pulses, as recently demonstrated at the W-band (95 GHz, ∼3.5 T). 8,9 In addition, high-field measurements open new opportunities for the use of metal ions as spin labels. The EPR spectra of highspin transition-metal ions with half-filled valence orbitals, such as Gd 3+ (S = 7/2) and Mn 2+ (S = 5/2), become much simpler at high fields, displaying an intense and relatively narrow central |−1/2⟩ → |1/2⟩ transition. 10,11 The width of this transition is proportional to D 2 /ν 0 , where D is the zero-field splitting (ZFS) parameter and ν 0 is the spectrometer frequency, leading to increased sensitivity with increased freque...

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“…This is unfortunately not possible with a single-mode resonator. A possible solution to that problem is the use of a tunable double-mode resonator (Tkach, Sicoli, H€ obartner, & Bennati, 2011) or by using a nonresonant sample holder (Cruickshank et al, 2009).…”

Section: G-band Peldor (High Magnetic Field)mentioning

confidence: 99%