Beriashvili, David, Ru Yao, Francesca D’Amico, Michaela Krafčíková, Andrei Gurinov, Adil Safeer, Xinyi Cai, et al. “A High-Field Cellular DNP-Supported Solid-State NMR Approach to Study Proteins with Sub-Cellular Specificity.” Chemical Science 14, no. 36 (2023): 9892–99.
https://doi.org/10.1039/D3SC02117C.
Studying the structural aspects of proteins within sub-cellular compartments is of growing interest. Dynamic nuclear polarization supported solid-state NMR (DNP-ssNMR) is uniquely suited to provide such information, but critically lacks the desired sensitivity and resolution. Here we utilize SNAPol-1, a novel biradical, to conduct DNP-ssNMR at high-magnetic fields (800 MHz/527 GHz) inside HeLa cells and isolated cell nuclei electroporated with [13C, 15N] labeled ubiquitin. We report that SNAPol-1 passively diffuses and homogenously distributes within whole cells and cell nuclei providing ubiquitin spectra of high sensitivity and remarkably improved spectral resolution. For cell nuclei, physical enrichment facilitates a further 4-fold decrease in measurement time and provides an exclusive structural view of the nuclear ubiquitin pool. Taken together, these advancements enable atomic interrogation of protein conformational plasticity at atomic resolution and with sub-cellular specificity.