Fig. 3. In-source dissociation causes linear dependence of the ligand-bound dsA2 fraction on cone voltage.

a Native MS fractions of dsA2 and dsA2/erucamide are likewise dependent on the activation energy of the cone voltage. As the cone voltage is raised, the percentage of empty dsA2 increases because the adduct binding cannot withstand the energy in the ion source and therefore dissociates. The proportionality is linear, and therefore a theoretical cone voltage of 48 V can be calculated at which a full occupancy with ligand would be reached. This voltage is too low to obtain stable electrospray and a resolved spectrum. b Peptide binding is equally subject to ISD. NV9-bound dsA2 (light blue and dark blue annotated peaks) become less prominent with increasing cone voltage, while empty dsA2 (yellow) increases at the same time.