title: W-band orientation selective DEER measurements on a Gd3+/nitroxide mixed-labeled protein dimer with a dual mode cavity.
abstract: Double electron-electron resonance (DEER) at W-band (95 GHz) was applied to measure the distance between a pair of nitroxide and Gd(3+) chelate spin labels, about 6 nm apart, in a homodimer of the protein ERp29. While high-field DEER measurements on systems with such mixed labels can be highly attractive in terms of sensitivity and the potential to access long distances, a major difficulty arises from the large frequency spacing (about 700 MHz) between the narrow, intense signal of the Gd(3+) central transition and the nitroxide signal. This is particularly problematic when using standard single-mode cavities. Here we show that a novel dual-mode cavity that matches this large frequency separation dramatically increases the sensitivity of DEER measurements, allowing evolution times as long as 12 μs in a protein. This opens the possibility of accessing distances of 8 nm and longer. In addition, orientation selection can be resolved and analyzed, thus providing additional structural information. In the case of W-band DEER on a Gd(3+)-nitroxide pair, only two angles and their distributions have to be determined, which is a much simpler problem to solve than the five angles and their distributions associated with two nitroxide spin labels.
authors: Department of Chemical Physics, Weizmann Institute of Science, Rehovot, Israel.
authors: Ilia
authors: I
authors: Kaminker
authors: Igor
authors: I
authors: Tkach
authors: Nurit
authors: N
authors: Manukovsky
authors: Thomas
authors: T
authors: Huber
authors: Hiromasa
authors: H
authors: Yagi
authors: Gottfried
authors: G
authors: Otting
authors: Marina
authors: M
authors: Bennati
authors: Daniella
authors: D
authors: Goldfarb
mesh_terms: D000465:Algorithms; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D005682:Gadolinium / Q000737:chemistry*; D009589:Nitrogen Oxides / Q000737:chemistry*; D011506:Proteins / Q000032:analysis / Q000737:chemistry*
chemical_list: D009589:Nitrogen Oxides; D011506:Proteins; D005682:Gadolinium; C039900:nitroxyl