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Solid-State NMR Spectra of Lipid-Anchored Proteins under Magic Angle Spinning
journal contribution
posted on 2014-03-06, 00:00 authored by Kaoru Nomura, Erisa Harada, Kenji Sugase, Keiko ShimamotoSolid-state NMR is a promising tool
for elucidating membrane-related
biological phenomena. We achieved the measurement of high-resolution
solid-state NMR spectra for a lipid-anchored protein embedded in lipid
bilayers under magic angle spinning (MAS). To date, solid-state NMR
measurements of lipid-anchored proteins have not been accomplished
due to the difficulty in supplying sufficient amount of stable isotope
labeled samples in the overexpression of lipid-anchored proteins requiring
complex posttranslational modification. We designed a pseudo lipid-anchored
protein in which the protein component was expressed in E. coli and attached to a chemically synthesized
lipid-anchor mimic. Using two types of membranes, liposomes and bicelles,
we demonstrated different types of insertion procedures for lipid-anchored
protein into membranes. In the liposome sample, we were able to observe
the cross-polarization and the 13C–13C chemical shift correlation spectra under MAS, indicating that the
liposome sample can be used to analyze molecular interactions using
dipolar-based NMR experiments. In contrast, the bicelle sample showed
sufficient quality of spectra through scalar-based experiments. The
relaxation times and protein–membrane interaction were capable
of being analyzed in the bicelle sample. These results demonstrated
the applicability of two types of sample system to elucidate the roles
of lipid-anchors in regulating diverse biological phenomena.