On-Cell Detection of Polysaccharide One-Bond ¹J_CH Couplings by Proton-Detected Solid-State NMR

The one-bond proton-carbon coupling constant (¹J_CH) is an insightful probe of carbohydrate configuration. Equatorial and axial protons at the C₁ position typically exhibit distinct ¹J_CH values, enabling NMR measurements to distinguish α- and β-configurations in carbohydrates. In principle, such measurements could provide insights into carbohydrates in the cell walls of intact microbes. However, traditionally, these measurements are performed by solution NMR with carbohydrates that were extracted, solubilized and fractionated, leaving the biological relevance of the measurements uncertain. Here, we demonstrate that ¹H-detected solid-state NMR with fast magic-angle spinning allows quantitative measurements of ¹J_CH couplings for mobile capsular polysaccharides, directly on submilligram amounts of pathogenic cells. Our approach is demonstrated on intact cells of the pathogenic yeast Cryptococcus neoformans. High-resolution proton-detected spectra enabled the determination of coupling constants for five mobile polysaccharide units of the cryptococcal capsule, revealing their native configurations and confirming previous solution NMR-based anomeric configuration assignments.