Chemical Derivatization Enables MALDI-TOF-Based High-Throughput Screening for Microbial Trimethylamine (TMA)-Lyase Inhibitors

MALDI-TOF-Based Affinity Selection Mass Spectrometry for Automated Screening of Protein–Ligand Interactions at High Throughput

“…Compared to ESI-based MS, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS provides the sampling speed required for HTS and has been established as a valuable readout strategy for large-scale screening campaigns against the catalytic activity of enzymatic targets. [13][14][15][16][17] The adoption of the technology to interrogate protein-ligand interaction has been the subject of a number of proof-of-principle studies using ultrafiltration or affinity capture-based workflows to isolate protein-ligand complexes from incubation mixtures. [18][19][20] Although these studies have demonstrated the general feasibility of using MALDI-TOF MS for label-free small-molecule detection, a report of its combination with an HTS-capable automation platform is unprecedented.…”

High-Throughput Mass Spectrometry for Hit Identification: Current Landscape and Future Perspectives

“…More recently, and building on their success, the same group reported on screening for inhibitors of TMA generating enzymes in gut bacteria. 76 They developed a MALDI-ToF assay for quantification of secreted TMA levels within bacterial cultures to assess microbial TMA-lyase activity. A key element of the assay was derivatization of the sample generated in microbial assay using biotin polyethylneoxide iodoacetamide (BPI) in order to increase the molecular weight of the analyte and thus avoid interference from matrix and other background ions.…”

MALDI-TOF Mass Spectrometry-Based High-Throughput Screening for Inhibitors of the Cytosolic DNA Sensor cGAS

“…[29][30][31][32][33] We have recently demonstrated the successful integration of fast direct MALDI-TOF MS into the HTS environment and its adaptation as a versatile and label-free readout strategy for in vitro enzyme assays. [34][35][36] In this work, we advance beyond the proof of concept and demonstrate its application in a large-scale fulldiversity HTS campaign against the catalytic activity of human cGAS. MS-based direct and label-free quantification of cGAMP was used to track compound-dependent enzyme inhibition at high speed.…”

“…29–33 We have recently demonstrated the successful integration of fast direct MALDI-TOF MS into the HTS environment and its adaptation as a versatile and label-free readout strategy for in vitro enzyme assays. 34–36…”