Nano-Mole Scale Side-Chain Signal Assignment by 1H-Detected Protein Solid-State NMR by Ultra-Fast Magic-Angle Spinning and Stereo-Array Isotope Labeling

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dc.contributor.authorWang, Songlin
dc.contributor.authorParthasarathy, Sudhakar
dc.contributor.authorNishiyama, Yusuke
dc.contributor.authorEndo, Yuki
dc.contributor.authorNemoto, Takahiro
dc.contributor.authorYamauchi, Kazuo
dc.contributor.authorAsakura, Tetsuo
dc.contributor.authorTakeda, Mitsuhiro
dc.contributor.authorTerauchi, Tsutomu
dc.contributor.authorKainosho, Masatsune
dc.contributor.authorIshii, Yoshitaka
dc.date.accessioned2020-05-04T10:26:11Z
dc.date.available2020-05-04T10:26:11Z
dc.date.issued2015-04
dc.description.abstractWe present a general approach in 1H-detected 13C solid-state NMR (SSNMR) for side-chain signal assignments of 10-50 nmol quantities of proteins using a combination of a high magnetic field, ultra-fast magic-angle spinning (MAS) at ~80 kHz, and stereo-array-isotope-labeled (SAIL) proteins [Kainosho M. et al., Nature 440, 52–57, 2006]. First, we demonstrate that 1H indirect detection improves the sensitivity and resolution of 13C SSNMR of SAIL proteins for side-chain assignments in the ultra-fast MAS condition. 1H-detected SSNMR was performed for micro-crystalline ubiquitin (~55 nmol or ~0.5mg) that was SAIL-labeled at seven isoleucine (Ile) residues. Sensitivity was dramatically improved by 1H-detected 2D 1H/13C SSNMR by factors of 5.4-9.7 and 2.1-5.0, respectively, over 13C-detected 2D 1H/13C SSNMR and 1D 13C CPMAS, demonstrating that 2D 1H-detected SSNMR offers not only additional resolution but also sensitivity advantage over 1D 13C detection for the first time. High 1H resolution for the SAIL-labeled side-chain residues offered reasonable resolution even in the 2D data. A 1H-detected 3D 13C/13C/1H experiment on SAIL-ubiquitin provided nearly complete 1H and 13C assignments for seven Ile residues only within ~2.5 h. The results demonstrate the feasibility of side-chain signal assignment in this approach for as little as 10 nmol of a protein sample within ~3 days. The approach is likely applicable to a variety of proteins of biological interest without any requirements of highly efficient protein expression systems.en_US
dc.identifier.citation: Wang S, Parthasarathy S, Nishiyama Y, Endo Y, Nemoto T, Yamauchi K, et al. (2015) NanoMole Scale Side-Chain Signal Assignment by 1 HDetected Protein Solid-State NMR by Ultra-Fast Magic-Angle Spinning and Stereo-Array Isotope Labeling. PLoS ONE 10(4): e0122714. doi:10.1371/ journal.pone.0122714en_US
dc.identifier.issn1932-6203
dc.identifier.other10.1371/journal.pone.0122714
dc.identifier.urihttps://journals.plos.org/plosone/article?id=10.1371/journal.pone.0122714
dc.identifier.urihttps://doi.org/10.1371/journal.pone.0122714
dc.identifier.urihttp://nur.nu.edu.kz/handle/123456789/4595
dc.language.isoenen_US
dc.publisherPublic Library of Scienceen_US
dc.relation.ispartofseriesPLoS ONE;
dc.rightsAttribution-NonCommercial-ShareAlike 3.0 United States*
dc.rights.urihttp://creativecommons.org/licenses/by-nc-sa/3.0/us/*
dc.subjectResearch Subject Categories::NATURAL SCIENCES::Chemistryen_US
dc.subjectnano-moleen_US
dc.subjectmagic-angle spinningen_US
dc.subjectMASen_US
dc.subjectstereo-array-isotope-labeleden_US
dc.subjectSAILen_US
dc.subjectprotein expression systemen_US
dc.titleNano-Mole Scale Side-Chain Signal Assignment by 1H-Detected Protein Solid-State NMR by Ultra-Fast Magic-Angle Spinning and Stereo-Array Isotope Labelingen_US
dc.typeArticleen_US
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