PMID- 28205016 OWN - NLM STAT- MEDLINE DCOM- 20171207 LR - 20181113 IS - 1573-5001 (Electronic) IS - 0925-2738 (Linking) VI - 67 IP - 2 DP - 2017 Feb TI - Sensitivity enhancement for membrane proteins reconstituted in parallel and perpendicular oriented bicelles obtained by using repetitive cross-polarization and membrane-incorporated free radicals. PG - 135-144 LID - 10.1007/s10858-017-0090-0 [doi] AB - Multidimensional separated local-field and spin-exchange experiments employed by oriented-sample solid-state NMR are essential for structure determination and spectroscopic assignment of membrane proteins reconstituted in macroscopically aligned lipid bilayers. However, these experiments typically require a large number of scans in order to establish interspin correlations. Here we have shown that a combination of optimized repetitive cross polarization (REP-CP) and membrane-embedded free radicals allows one to enhance the signal-to-noise ratio by factors 2.4-3.0 in the case of Pf1 coat protein reconstituted in magnetically aligned bicelles with their normals being either parallel or perpendicular to the main magnetic field. Notably, spectral resolution is not affected at the 2:1 radical-to-protein ratio. Spectroscopic assignment of Pf1 coat protein in the parallel bicelles has been established as an illustration of the method. The proposed methodology will advance applications of oriented-sample NMR technique when applied to samples containing smaller quantities of proteins and three-dimensional experiments. FAU - Koroloff, Sophie N AU - Koroloff SN AD - Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204, USA. FAU - Tesch, Deanna M AU - Tesch DM AD - Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204, USA. AD - Shaw University, 118 E. South Street, Raleigh, NC, 27601, USA. FAU - Awosanya, Emmanuel O AU - Awosanya EO AD - Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204, USA. FAU - Nevzorov, Alexander A AU - Nevzorov AA AUID- ORCID: 0000-0001-9676-6139 AD - Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC, 27695-8204, USA. alex_nevzorov@ncsu.edu. LA - eng PT - Journal Article PT - Research Support, U.S. Gov't, Non-P.H.S. DEP - 20170215 PL - Netherlands TA - J Biomol NMR JT - Journal of biomolecular NMR JID - 9110829 RN - 0 (Free Radicals) RN - 0 (Membrane Proteins) RN - 0 (Viral Proteins) SB - IM MH - Bacteriophage Pf1 MH - Free Radicals/*chemistry MH - *Magnetic Resonance Spectroscopy/methods/standards MH - Membrane Proteins/*chemistry MH - Signal-To-Noise Ratio MH - Viral Proteins/chemistry OTO - NOTNLM OT - Magnetically aligned bicelles OT - Membrane proteins OT - Oriented-sample NMR (OS-NMR) OT - Paramagnetic relaxation enhancement OT - Pf1 OT - Repetitive cross-polarization OT - Sensitivity enhancement OT - Spectroscopic assignment EDAT- 2017/02/17 06:00 MHDA- 2017/12/08 06:00 CRDT- 2017/02/17 06:00 PHST- 2016/12/01 00:00 [received] PHST- 2017/01/19 00:00 [accepted] PHST- 2017/02/17 06:00 [pubmed] PHST- 2017/12/08 06:00 [medline] PHST- 2017/02/17 06:00 [entrez] AID - 10.1007/s10858-017-0090-0 [pii] AID - 10.1007/s10858-017-0090-0 [doi] PST - ppublish SO - J Biomol NMR. 2017 Feb;67(2):135-144. doi: 10.1007/s10858-017-0090-0. Epub 2017 Feb 15.