Direct observation of glycans bonded to proteins and lipids at single molecule level

Proteins and lipids decorated with glycans are found throughout biological entities, playing roles in biological functions and dysfunctions. Current analytical strategies for these glycan-decorated biomolecules, termed glycoconjugates, rely on ensemble averaged methods that do not provide a full view of positions and structures of glycans attached at individual sites in a given molecule, especially for glycoproteins. Here we show single molecule analysis of glycoconjugates by direct imaging of individual glycoconjugate molecules using low-temperature scanning tunneling microscopy. Intact glycoconjugate ions from electrospray are soft-landed on surface for their direct single molecule imaging. The sub-molecular imaging resolution corroborated by quantum mechanical modeling unveils the entire structures and attachment sites of glycans in glycopeptides, glycolipids, N-glycoproteins, and O-glycoproteins densely decorated with glycans.