摘要:SummaryC-Mannosylation is a relatively rare form of protein glycosylation involving the attachment of an α-mannopyranosyl residue to C-2 of the indole moiety of the amino acid tryptophan. This type of linkage was initially discovered in RNase 2 from human urine but later confirmed to be present in many other important proteins. Based on NMR experiments and extensive molecular dynamics simulations on the hundred microsecond timescale we demonstrate that, for isolated glycopeptides and denatured RNase 2, the C-linked mannopyranosyl residue exists as an ensemble of conformations, among which1C4is the most abundant. However, for native RNase 2, molecular dynamics and NMR studies revealed that the mannopyranosyl residue favors a specific conformation, which optimally stabilizes the protein fold through a network of hydrogen bonds and which leads to a significant reduction of the protein dynamics on the microsecond timescale. Our findings contribute to the understanding of the biological role ofC-mannosylation.Graphical AbstractDisplay OmittedHighlights•NMR and MD show thatC-linked mannose exists as an ensemble of conformations•Conformation of mannose is influenced by the protein environment and solvent•In RNase 2 mannose favors a conformation that optimally stabilizes the protein fold•Efficient methods for analysis of a large number of MD trajectories are presentedBiochemistry; Structural Biology; Protein Structure Aspects