MS-TAFI: A Tool for the Analysis of Fragment Ions Generated from Intact Proteins

Abstract

Tandem mass spectrometry (MS/MS) spectra of intact proteins can be difficult to interpret owing to the variety of fragment ion types and abundances. This information is crucial for maximizing the information derived from top-down mass spectrometry of proteins and protein complexes. MS-TAFI (Mass Spectrometry Tool for the Analysis of Fragment Ions) is a free Pythonbased program which offers a streamlined approach to the data analysis and visualization of deconvoluted MS/MS data of intact proteins. The application also contains tools for native mass spectrometry experiments with the ability to search for fragment ions that retain ligands (holo ions) as well as visualize the location of charge sites obtained from 193 nm ultraviolet photodissociation data. The source code and complete application for MS-TAFI is available for download at https:// github.com/kylejuetten.

Top-Down Analysis of Supercharged Proteins Using Collision-, Electron-, and Photon-Based Activation Methods

“…Deconvoluted fragment ion abundances were analyzed with a custom-built Python program, MS-TAFI, to generate fragment abundance maps. 56 All ten UVPD fragment ion types (a, a+1, b, c, x, x+1, y, y-1, y-2, and z) were considered for sequence coverage and general analysis of fragment abundances. When evaluating preferential cleavage sites, each interresidue position is counted as one backbone cleavage site regardless of whether a,b,c or x,y,z ions are generated from cleavage of that inter-residue position.…”

Top-Down Analysis of Supercharged Proteins Using Collision-, Electron-, and Photon-Based Activation Methods

“…Fragments were identified using a 10 ppm error tolerance. Deconvoluted fragment ion abundances were analyzed with a custom-built Python program, MS-TAFI (), to generate fragment abundance maps . All ten UVPD fragment ion types ( a , a +1, b , c , x , x +1, y , y –1, y –2, and z ) were considered for sequence coverage and general analysis of fragment abundances.…”

“…To further examine the impacts of supercharging on preferential cleavages, backbone cleavage maps were created using a Python program, MS-TAFI . This program utilizes deconvoluted fragment ion abundances to display the abundances of fragments (normalized to the total fragment ion current) according to the backbone cleavage position from which they originated, allowing visualization of the pattern of backbone cleavages (“cut sites”) throughout the protein sequence.…”

“…Deconvoluted fragment ion abundances were analyzed with a custom-built Python program, MS-TAFI (https://github.com/kylejuetten), to generate fragment abundance maps. 56 All ten UVPD fragment ion types (a, a+1, b, c, x, x+1, y, y−1, y−2, and z) were considered for sequence coverage and general analysis of fragment abundances. When evaluating preferential cleavage sites, each inter-residue position is counted as one backbone cleavage site regardless of whether a,b,c or x,y,z ions are generated from cleavage of that inter-residue position.…”

“…To further examine the impacts of supercharging on preferential cleavages, backbone cleavage maps were created using a Python program, MS-TAFI. 56 This program utilizes deconvoluted fragment ion abundances to display the abundances of fragments (normalized to the total fragment ion current) according to the backbone cleavage position from which they originated, allowing visualization of the pattern of backbone cleavages ("cut sites") throughout the protein sequence. This format graphically illustrates the impact of highly preferred cleavage sites as a function of charge state and also improves visualization of variations in coverage of the protein sequence for each activation method.…”