Glycans have several elements that contribute to their structural complexity, involving a range of monosaccharide building blocks, configuration of linkages between residues and various degrees of branching on a given structure. Their analysis remains challenging and resolving minor isomeric variants can be difficult, in particular terminal fucosylated Lewis and blood group antigens present on N- and O-glycans. Accurately characterizing these isomeric structures by current techniques is not straightforward and typically requires a combination of methods and/or sample derivatization. Yet the ability to monitor the occurrence of these epitopes is important as structural changes are associated with several human diseases. The use of ion mobility-mass spectrometry (IM-MS), which separates ions in the gas phase based on their size, charge and shape, offers a new potential tool for glycan analysis and recent reports have demonstrated its potential for glycomics. Here we show that Lewis and blood group isomers, which have identical fragmentation spectra, exhibit very distinctive IM drift times and collision cross sections (CCS). We show that IM-MS/MS analysis can rapidly and accurately differentiate epitopes from parotid gland N-glycans and milk oligosaccharides based on fucosylated fragment ions with characteristic CCSs.

Although there have been substantial improvements in glycan analysis over the past decade, the lack of both high-resolution and high-throughput methods hampers progress in glycomics.

A vital step in HIV vaccine development strategies has been the observation that some infected individuals generate broadly neutralizing antibodies that target the glycans on the surface of HIV-1 gp120. These antibodies target glycan epitopes on viral envelope spikes and yet the positions and degree of occupancy of glycosylation sites is diverse. Therefore, there is a need to understand glycosylation occupancy on recombinant immunogens. The sheer number of potential glycosylation sites and degree of chemical heterogeneity impedes assessing the global sequon occupancy of gp120 glycoforms. Here, we trap the glycan processing of recombinant gp120 to generate homogenous glycoforms, facilitating occupancy assessment by intact mass spectrometry. We show that gp120 monomers of the BG505 strain contain either fully occupied sequons or missing one and sometimes two glycans across the molecule. This biosynthetic engineering approach enables the analysis of therapeutically important glycoproteins otherwise recalcitrant to analysis by native mass spectrometry.

Bloodgroup oligosaccharides have been derivatized with labels common in HPLC and evaluated regarding their ion mobility behaviour.

The combination of 2,2':6',2''-terpyridines (tpy) and Ru II is knownt od eliverm olecular and supramolecular assembliesw ith remarkablep roperties.H ere new Ru II complexes, with modified tpy ligandss ubstituted with varying numbers of dimethlyamino groups,a re presented. Electrochemistry shows that the incorporation of the strongly electron-donatingg roups on the tpy ligands leads to an egative shift of the Ru II oxidation potentialb yc lose to 1V .T he reductive electrochemical responses are strongly dependent on the nature of the working electrode, with glassyc arbon and gold working electrodes showing the best results. These observations led to the development of am odified Optically Transparent Thin Layer Electrochemical (OTTLE)c ell, based on ag old working electrode. The use of UV/Vis/NIR spectroelectrochemical methods with that OTTLE cell, together with simulations of the cyclic voltammograms, allowed the characterization of four reduction steps in these complexes, the final two of which lead to bond activationsa tt he ruthenium center.T his observation is to the best of our knowledge unprecedented in coordinatively saturated complexes of type [Ru (tpy) 2 ] 2 + .T he variousr edox states of the complexes were characterized by EPR spectroelectrochemistry and through DFT calculations. The results presented here establish these substitutedt py ligands as highly attractive ligands in coordination chemistry,a nd displayt he utility of ag old-basedO TTLE cell for spectroelectrochemicalm easurements.