Ultra-high-field solid-state NMR of intact Saccharomyces cerevisiae cells at 1.2 GHz

The increasing availability of ultra-high-field NMR spectrometers is opening new opportunities for the characterization of complex and structurally heterogeneous biological assemblies by solid-state NMR, including lipid membranes, cell walls, and even intact cells. Here, we investigate the use of a 1.2 GHz (28.2 T) spectrometer, the highest magnetic field currently commercially available, for the analysis of intact Saccharomyces cerevisiae cells. Budding yeast is an experimentally tractable eukaryotic model organism that is widely used in biotechnology and industrial fermentation. Characterization of their cell-wall polysaccharides is essential for understanding cell integrity, stress responses, and strain-dependent functional properties. We compared one-dimensional and two-dimensional carbon-detected solid-state NMR experiments recorded at 1.2 GHz and 600 MHz under slow magic-angle spinning (11 kHz) conditions to evaluate gains in spectral resolution. Across representative polysaccharide resonances, linewidths in ppm are reduced by approximately 30-35% at 1.2 GHz. In addition, proton-detected experiments performed at 1.2 GHz under fast magic-angle spinning (100 kHz) enabled the acquisition of highly resolved 13 C- 1 H correlation spectra that probe both rigid and mobile polysaccharide components of the cell wall. Proton detection further revealed site-specific linewidth variations of 0.22-0.38 ppm for rigid polysaccharides and as low as 0.043 ppm for mobile polysaccharides, enabling the identification of structurally distinct polysaccharide forms that are not resolved by direct 13 C detection. Overall, these results demonstrate that ultra-high-field solid-state NMR substantially enhances spectral resolution by increasing chemical-shift dispersion and revealing hidden structural heterogeneity, thereby enabling detailed characterization of intact yeast cell-wall architecture.