PepFoot: A Software Package for Semiautomated Processing of Protein Footprinting Data

Covalent
footprinting of proteins using reactive intermediates
such as radicals and carbenes is emerging as a valuable tool for mapping
surface accessibility, and hence binding sites of proteins. The approach
generates a significant amount of mass spectrometry (MS) data, which
can be time-consuming to process manually. PepFoot, a software package
that allows semiautomated processing of MS data from footprinting
experiments, is described. By using the open source .mz5 file format,
it is able to accept data from all the major instrument manufacturers.
Following manual user interrogation of one data file within a user-friendly
GUI, the software then automates determination of the degree of fractional
modification (fm) with the footprinting
agent across a batch of experimental data. This greatly increases
efficiency and throughput compared to manual analysis of each file,
and provides initial scrutiny and confidence compared to fully automated
analysis. Histogram plots of fm for each
peptide from the footprinted protein may be displayed within PepFoot
and mapped onto an imported protein structure to reveal differential
labeling patterns and hence binding sites. The software has been tested
on data from carbene and hydroxyl radical labeling experiments to
demonstrate its broad utility. PepFoot is released under the LGPL
version 3 license, and is available for Windows, MacOS, and Linux
systems at github.com/jbellamycarter/pepfoot.