The Utilization of the Search Engine, Bolt, to Decrease Search Time and Increase Peptide Identifications in Hydroxyl Radical Protein Footprinting-based Workflows

Hydroxyl radical protein footprinting (HRPF) utilizes hydroxyl radicals to covalently modify solvent exposed regions of proteins. When coupled with mass spectrometry, HRPF can provide insightful information on protein structural changes including inside cells. However, the complex mixture of proteins and modifications makes identification a complicated task. To search all of the HRPF-induced modification combinations across the full proteome, requires substantial computational power and still can take days to search. To drastically decrease processing time and improve identifications, a novel cloud-based search engine, Bolt, was used to search for HRPF modifications in comparison to a commonly used search engine, Sequest. A 35% increase in the identification of modified peptides was observed in Bolt compared to Sequest with a decrease in computation time.