Glykan-Sequenzierung an Oberflächen – Kombination von präparativer Massenspektrometrie und Rastersondenmikroskopie zur Entschlüsselung des Zuckercodes
Zusammenfassung der Projektergebnisse
This project has combined electrospray deposition and preparative mass spectrometry with scanning probe microscopy to achieve real-space imaging of glycans at the single-molecule level, and to determine the structure of these complex biomolecules through the highresolution images obtained by this method. Our results have shown single glycan molecules with atomic-scale detail, revealing not only their overall shape, but also the arrangement of functional groups and atoms inside the molecules, as well as the structural variations between individual glycans. In case of β-cyclodextrin, the location of intramolecular hydrogen bond networks – important for determining the conformation of the molecule – could be deduced from the atomically resolved images, and the combination of experiments with first principles calculations allowed for unraveling the atomic structure of the molecule. We have also employed the combination of electrospray deposition and scanning probe microscopy to study the sulfation pattern of glycosaminoglycans, a class of sulfated glycans that play important roles in cell surface biology in animals. The images have shown full-length glycosaminoglycan chains unfolded on an atomically flat surface, and revealed minute structural features that may enable the localization of the important sulfate modification in the molecules. In addition to deciphering the structure of glycans, we have successfully applied the method developed within this project for the imaging of other biomolecules on surfaces, including individual proteins and DNA molecules. While high-resolution single-biomolecule imaging enabled by preparative mass spectrometry is still confined to fundamental academic research, requiring custom-built instrumentation, the results herein have demonstrated its potential to advance the analysis of glycans and other biomolecules in the future.
Projektbezogene Publikationen (Auswahl)
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Atomically resolved imaging of the conformations and adsorption geometries of individual β-cyclodextrins with non-contact AFM. Nature Communications, 15(1).
Grabarics, Márkó; Mallada, Benjamín; Edalatmanesh, Shayan; Jiménez-Martín, Alejandro; Pykal, Martin; Ondráček, Martin; Kührová, Petra; Struwe, Weston B.; Banáš, Pavel; Rauschenbach, Stephan; Jelínek, Pavel & de, la Torre Bruno
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Cryo-EM of soft-landed β-galactosidase: Gas-phase and native structures are remarkably similar. Science Advances, 10(7).
Esser, Tim K.; Böhning, Jan; Önür, Alpcan; Chinthapalli, Dinesh K.; Eriksson, Lukas; Grabarics, Marko; Fremdling, Paul; Konijnenberg, Albert; Makarov, Alexander; Botman, Aurelien; Peter, Christine; Benesch, Justin L. P.; Robinson, Carol V.; Gault, Joseph; Baker, Lindsay; Bharat, Tanmay A. M. & Rauschenbach, Stephan
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High-resolution Imaging of Glycans by Scanning Tunnelling Microscopy Enabled by Electrospray Ion Beam Deposition (ESIBD). Glycoprotein Analysis, 329-354. Royal Society of Chemistry.
Rauschenbach, Stephan; Grabarics, Marko; Delbianco, Martina; Cortes, Juan; Schön, Christian; Tarrat, Nathalie; Wu, Xu & Anggara, Kelvin
