Top-Down MALDI-In-Source Decay-FTICR Mass Spectrometry of Isotopically Resolved Proteins

An accurate mass Measurement of a known protein provides information on potential amino acid deletions and post-translational modifications. Although this field is dominated by strategies based on electrospray ionization, Mass spectrometry (MS) methods using matrix-assisted laser desorption/ionization (MALDI) have the advantage of yielding predominantly singly charged precursor ions, thus avoiding peak overlap from different charge states of multiple species. Such MALDI-MS methods require mass measurement at ultrahigh resolution, which is provided by Fourier transform ion cyclotron resonance (FTICR) mass analyzers. Recently, using a MALDI-FTICR-MS platform equipped with a 15 T magnet, we reported on the mass analysis of intact human serum peptides and small proteins with isotopic resolution up to similar to 15 kDa and identified new proteoforms from an accurate measurement of mass distances. In the current study, we have used this FTICR system after an upgrade with a novel dynamically harmonized ICR cell, i.e., ParaCell, for mapping isotopically resolved intact proteins up to about 17 kDa and performed topdown MALDI in-source decay (ISD) analysis. Standard proteins myoglobin (m/z-value 16 950) and ribonuclease B (m/z-value 14 900) were measured with resolving powers of 62 000 and 61 000, respectively. Furthermore, it will be shown that (singly charged) MALDI-ISD fragment ions can be measured at isotopic resolution up to m/z-value 12 000 (e.g., resolving power 39 000 at m/z-value 12 000) providing more reliable identifications. Moreover, examples are presented of pseudo-MS3 experiments on ISD fragment ions from RNase B by collisional-induced dissociation (CID).