TOPP: A Novel Nitroxide‐Labeled Amino Acid for EPR Distance Measurements

Stoller, Sven; Sicoli, Giuseppe; Baranova, T. Yu.; Bennati, Marina; Diederichsen, Ulf

doi:10.1002/anie.201103315

Cited by 36 publications

(33 citation statements)

References 22 publications

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“…The ratios of the integrated bands associated with the H‐bonded vs free NH groups point clearly to almost fully developed (≥ 90%) 3 10 ‐helical structures for all peptides examined . These findings are in full agreement with the spectroscopic and X‐ray diffraction literature data of peptides rich in the known helicogenic TOAC and Aib residues in CDCl 3 solution and in the crystal state.…”

Section: Resultssupporting

confidence: 87%

“…More specifically, we analyze a series of peptides consisting of stretches of the non‐coded, host α‐amino acid α‐aminoisobutyric acid (Aib), combined with one or two 4‐amino‐1‐oxyl‐2,2,6,6‐tetramethylpiperidine‐4‐carboxylic acid (TOAC) guest residues. Both TOAC and Aib are known to strongly stabilize regular 3 10 ‐helices . The backbone and side‐chain conformations of TOAC are remarkably constrained, providing well‐defined distances between the stable nitroxide free radicals of two TOAC residues.…”

Section: Introductionmentioning

confidence: 99%

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Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances

et al. 2014

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For 3D-structure determination in biophysical systems EPR is rapidly gaining ground. Proteins labeled specifically with two nitroxide spin labels can be prepared, and several EPR methods are available for distance determination, which makes it possible to determine distance constraints. However, such methods require frozen solutions, potentially causing non-physiological states of the sample. Here, we target spin- spin interaction in liquid solution at room temperature using rigid model compounds. A series of 310 -helical peptides, based on α-aminoisobutyric acid (Aib), is synthesized with pairs of spin labels separated by three, four, and five amino acids. To avoid flexibility, the noncoded nitroxyl-containing α-amino acid TOAC that is rigidly connected with the peptide backbone, is used. The EPR spectra of the peptides show a decreasing amount of coupling between the two spin labels within this series. We suggest through-bond interaction as the dominating mechanism for exchange interaction (J) and find a stronger J-coupling than in the corresponding Ala-based TOAC-peptides investigated previously (Hanson, et al., J Am Chem Soc 1996, 118, 7618-7625). We speculate that stronger coupling in Aib- vs Ala- peptides is due to intrinsically stronger through-bond interaction in the Aib-based peptides.

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Section: Resultssupporting

confidence: 87%

Section: Introductionmentioning

confidence: 99%

Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances

et al. 2014

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“…Crystal structures of RX (PDB entry 3L2X) and the 4-phenyl analog of R1p (PDB entry 1ZUR) in T4 lysozyme show resolved electron densities for the entire side chain, unlike those for R1 which are resolved only to the disulfide due to internal disorder. The side chain TOPP (Figure 1d) [24] provides a fixed position of the spin in a protein, but so far is introduced by total synthesis, restricting its practical use to peptides or small proteins.…”

Section: Nitroxide Side Chainsmentioning

confidence: 99%

Technological advances in site-directed spin labeling of proteins

Hubbell

López

Altenbach

et al. 2013

Current Opinion in Structural Biology

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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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“…Thus, TOAC is liable to modify the secondary structure, for it is a helicogenic amino acid. Recently, the chiral unnatural amino acid 4-(3,3,5,5-tetramethyl-2,6-dioxo-4-oxylpiperazin-1-yl)-L-phenylglycine (TOPP, Scheme1) was introduced to overcome this problem [30]. The TOPP label allows for rotations about the C  -C  bond; however, this bond is collinear with the nitroxide N-O bond and the distance between the C  atom and the site of the electron spin lies within a narrow range.…”

Section: Labeling Strategy and Common Spin Labelsmentioning

confidence: 99%

Conformational dynamics and distribution of nitroxide spin labels

Jeschke

2013

Progress in Nuclear Magnetic Resonance Spectroscopy

136

115

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Long-range distance measurements based on paramagnetic relaxation enhancement (PRE) in NMR, quantification of surface water dynamics near biomacromolecules by Overhauser dynamic nuclear polarization (DNP) and sensitivity enhancement by solid-state DNP all depend on introducing paramagnetic species into an otherwise diamagnetic NMR sample. The species can be introduced by site-directed spin labeling, which offers precise control for positioning the label in the sequence of a biopolymer. However, internal flexibility of the spin label gives rise to dynamic processes that potentially influence PRE and DNP behavior and leads to a spatial distribution of the electron spin even in solid samples. Internal dynamics of spin labels and their static conformational distributions have been studied mainly by electron paramagnetic resonance spectroscopy and molecular dynamics simulations, with a large body of results for the most widely applied methanethiosulfonate spin label MTSL. These results are critically discussed in a unifying picture based on rotameric states of the group that carries the spin label. Deficiencies in our current understanding of dynamics and conformations of spin labeled groups and of their influence on NMR observables are highlighted and directions for further research suggested.

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TOPP: A Novel Nitroxide‐Labeled Amino Acid for EPR Distance Measurements

Cited by 36 publications

References 22 publications

Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances

Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances

Technological advances in site-directed spin labeling of proteins

Conformational dynamics and distribution of nitroxide spin labels

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TOPP: A Novel Nitroxide‐Labeled Amino Acid for EPR Distance Measurements

Cited by 36 publications

References 22 publications

Conformation and EPR characterization of rigid, 310‐helical peptides with TOAC spin labels: Models for short distances

Conformation and EPR characterization of rigid, 310‐helical peptides with TOAC spin labels: Models for short distances

Technological advances in site-directed spin labeling of proteins

Conformational dynamics and distribution of nitroxide spin labels

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Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances

Conformation and EPR characterization of rigid, 3₁₀‐helical peptides with TOAC spin labels: Models for short distances