1H/13C/15N triple-resonance experiments for structure determinaton of membrane proteins by oriented-sample NMR.
 

1H/13C/15N triple-resonance experiments for structure determinaton of membrane proteins by oriented-sample NMR.

http://www.bionmr.com//www.ncbi.nlm....PubMedLink.gif Related Articles 1H/13C/15N triple-resonance experiments for structure determinaton of membrane proteins by oriented-sample NMR.

Solid State Nucl Magn Reson. 2020 Nov 24;111:101701

Authors: Lapin J, Awosanya EO, Esteves RJA, Nevzorov AA

Abstract
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*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.


PMID: 33260039 [PubMed - as supplied by publisher]



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