A theoretical assessment of structure determination of multi-span membrane proteins by oriented sample solid-state NMR spectroscopy

Oriented sample solid-state NMR (OS-ssNMR) spectroscopy allows the direct determination of the structure and topology of membrane proteins reconstituted into aligned lipid bilayers. Although OS-ssNMR theoretically has no upper size limit, its application to multi-span membrane proteins has not been established because most studies have been restricted to single- or dual-span proteins and peptides. Here, we present a critical assessment of the application of this method to multi-span membrane proteins. We used molecular dynamics simulations to back-calculate [¹⁵N-¹H] separated local field (SLF) spectra from a G protein-coupled receptor (GPCR) and show that fully resolved spectra can be obtained theoretically for a multi-span membrane protein with currently achievable resonance linewidths.