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

Handle URI:
http://hdl.handle.net/10754/550050
Title:
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
Authors:
Wang, Songlin; Parthasarathy, Sudhakar; Nishiyama, Yusuke; Endo, Yuki; Nemoto, Takahiro; Yamauchi, Kazuo; Asakura, Tetsuo; Takeda, Mitsuhiro; Terauchi, Tsutomu; Kainosho, Masatsune; Ishii, Yoshitaka
Abstract:
We 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.
KAUST Department:
Nuclear Magnetic Resonance Core Lab
Citation:
Wang S, Parthasarathy S, Nishiyama Y, Endo Y, Nemoto T, et al. (2015) 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. PLoS ONE 10(4): e0122714. doi:10.1371/journal.pone.0122714
Journal:
PLOS ONE
Issue Date:
9-Apr-2015
DOI:
10.1371/journal.pone.0122714
PubMed ID:
25856081
PubMed Central ID:
PMC4391754
Type:
Article
ISSN:
1932-6203
Additional Links:
http://dx.plos.org/10.1371/journal.pone.0122714
Appears in Collections:
Articles

Full metadata record

DC FieldValue Language
dc.contributor.authorWang, Songlinen
dc.contributor.authorParthasarathy, Sudhakaren
dc.contributor.authorNishiyama, Yusukeen
dc.contributor.authorEndo, Yukien
dc.contributor.authorNemoto, Takahiroen
dc.contributor.authorYamauchi, Kazuoen
dc.contributor.authorAsakura, Tetsuoen
dc.contributor.authorTakeda, Mitsuhiroen
dc.contributor.authorTerauchi, Tsutomuen
dc.contributor.authorKainosho, Masatsuneen
dc.contributor.authorIshii, Yoshitakaen
dc.date.accessioned2015-04-14T07:53:25Zen
dc.date.available2015-04-14T07:53:25Zen
dc.date.issued2015-04-09en
dc.identifier.citationWang S, Parthasarathy S, Nishiyama Y, Endo Y, Nemoto T, et al. (2015) 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. PLoS ONE 10(4): e0122714. doi:10.1371/journal.pone.0122714en
dc.identifier.issn1932-6203en
dc.identifier.pmid25856081en
dc.identifier.doi10.1371/journal.pone.0122714en
dc.identifier.urihttp://hdl.handle.net/10754/550050en
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
dc.relation.urlhttp://dx.plos.org/10.1371/journal.pone.0122714en
dc.rightsThis is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are crediteden
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
dc.typeArticleen
dc.contributor.departmentNuclear Magnetic Resonance Core Laben
dc.identifier.journalPLOS ONEen
dc.identifier.pmcidPMC4391754en
dc.eprint.versionPublisher's Version/PDFen
dc.contributor.institutionDepartment of Chemistry and University of Illinois at Chicago, Chicago, Illinois, United States of Americaen
dc.contributor.institutionRIKEN CLST-JEOL collaboration center, RIKEN, Yokohama, Kanagawa, Japanen
dc.contributor.institutionSchool of Science and Technology, Nazarbayev University, Astana, Kazakhstanen
dc.contributor.institutionDepartment of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo, Japanen
dc.contributor.institutionJEOL RESONANCE Inc., Akishima, Tokyo, Japanen
dc.contributor.institutionStructural Biology Research Center, Graduate School of Science, Furocho, Chikusa-ku, Nagoya University, Nagoya, Japan 464–8601en
dc.contributor.institutionSAIL Technologies Co., Inc., Tsurumi-ku, Yokohama, Kanagawa, Japanen
dc.contributor.institutionCenter for Priority Areas, Tokyo Metropolitan University, Tokyo, Japanen
dc.contributor.institutionCenter for Structural Biology, University of Illinois at Chicago, Chicago, Illinois, United States of Americaen
kaust.authorYamauchi, Kazuoen

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