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High resolving power ion mobility (IM) allows for accurate characterization of complex mixtures in high-throughput IM mass spectrometry (IM-MS) experiments. We previously demonstrated that pure component IM-MS data can be extracted from IM unresolved post-IM/collision-induced dissociation (CID) MS data using automated ion mobility deconvolution (AIMD) software [Matthew Brantley, Behrooz Zekavat, Brett Harper, Rachel Mason, and Touradj Solouki, J. Am. Soc. Mass Spectrom. , 2014, 25 , 1810–1819]. In our previous reports, we utilized a quadrupole ion filter for m / z -isolation of IM unresolved monoisotopic species prior to post-IM/CID MS. Here, we utilize a broadband IM-MS deconvolution strategy to remove the m / z -isolation requirement for successful deconvolution of IM unresolved peaks. Broadband data collection has throughput and multiplexing advantages; hence, elimination of the ion isolation step reduces experimental run times and thus expands the applicability of AIMD to high-throughput bottom-up proteomics. We demonstrate broadband IM-MS deconvolution of two separate and unrelated pairs of IM unresolved isomers ( viz. , a pair of isomeric hexapeptides and a pair of isomeric trisaccharides) in a simulated complex mixture. Moreover, we show that broadband IM-MS deconvolution improves high-throughput bottom-up characterization of a proteolytic digest of rat brain tissue. To our knowledge, this manuscript is the first to report successful deconvolution of pure component IM and MS data from an IM-assisted data-independent analysis (DIA) or HDMS E dataset. 

Abstract Repulsive electrostatic forces between prion‐like proteins are a barrier against aggregation. In neuropharmacology, however, a prion's net charge (Z) is not a targeted parameter. Compounds that selectively boost prionZremain unreported. Here, we synthesized compounds that amplified the negative charge of misfolded superoxide dismutase‐1 (SOD1) by acetylating lysine‐NH₃⁺in amyloid‐SOD1, without acetylating native‐SOD1. Compounds resembled a “ball and chain” mace: a rigid amyloid‐binding “handle” (benzothiazole, stilbene, or styrylpyridine); an aryl ester “ball”; and a triethylene glycol chain connecting ball to handle. At stoichiometric excess, compounds acetylated up to 9 of 11 lysine per misfolded subunit (ΔZ_fibril=−8100 per 10³subunits). Acetylated amyloid‐SOD1 seeded aggregation more slowly than unacetylated amyloid‐SOD1 in vitro and organotypic spinal cord (these effects were partially due to compound binding). Compounds exhibited reactivity with other amyloid and non‐amyloid proteins (e.g., fibrillar α‐synuclein was peracetylated; serum albumin was partially acetylated; carbonic anhydrase was largely unacetylated).