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1H/13C/15N triple-resonance experiments for structure determinaton of membrane proteins by oriented-sample NMR.
Lapin, Joel; Awosanya, Emmanuel O; Esteves, Richard J A; Nevzorov, Alexander A.
Afiliación
  • Lapin J; Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204 USA.
  • Awosanya EO; Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204 USA.
  • Esteves RJA; Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204 USA.
  • Nevzorov AA; Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204 USA. Electronic address: alex_nevzorov@ncsu.edu.
Solid State Nucl Magn Reson ; 111: 101701, 2021 02.
Article en En | MEDLINE | ID: mdl-33260039
The benefits of triple-resonance experiments for structure determination of macroscopically oriented membrane proteins by solid-state NMR are discussed. While double-resonance 1H/15N experiments are effective for structure elucidation of alpha-helical domains, extension of the method of oriented samples to more complex topologies and assessing side-chain conformations necessitates further development of triple-resonance (1H/13C/15N) NMR pulse sequences. Incorporating additional spectroscopic dimensions involving 13C spin-bearing nuclei, however, introduces essential complications arising from the wide frequency range of the 1H-13C dipolar couplings and 13C CSA (>20 â€‹kHz), and the presence of the 13C-13C homonuclear dipole-dipole interactions. The recently reported ROULETTE-CAHA pulse sequence, in combination with the selective z-filtering, can be used to evolve the structurally informative 1H-13C dipolar coupling arising from the aliphatic carbons while suppressing the signals from the carbonyl and methyl regions. Proton-mediated magnetization transfer under mismatched Hartman-Hahn conditions (MMHH) can be used to correlate 13C and 15N nuclei in such triple-resonance experiments for the subsequent 15N detection. The recently developed pulse sequences are illustrated for n-acetyl Leucine (NAL) single crystal and doubly labeled Pf1 coat protein reconstituted in magnetically aligned bicelles. An interesting observation is that in the case of 15N-labeled NAL measured at 13C natural abundance, the triple (1H/13C/15N) MMHH scheme predominantly gives rise to long-range intermolecular magnetization transfers from 13C to 15N spins; whereas direct Hartmann-Hahn 13C/15N transfer is entirely intramolecular. The presented developments advance NMR of oriented samples for structure determination of membrane proteins and liquid crystals.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Protones / Proteínas de la Membrana Idioma: En Revista: Solid State Nucl Magn Reson Asunto de la revista: DIAGNOSTICO POR IMAGEM / MEDICINA NUCLEAR Año: 2021 Tipo del documento: Article Pais de publicación: Países Bajos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Protones / Proteínas de la Membrana Idioma: En Revista: Solid State Nucl Magn Reson Asunto de la revista: DIAGNOSTICO POR IMAGEM / MEDICINA NUCLEAR Año: 2021 Tipo del documento: Article Pais de publicación: Países Bajos