PepShell: Visualization of Conformational Proteomics Data

Abstract

Proteins are dynamic molecules; they undergo crucial conformational changes induced by post-translational modifications and by binding of cofactors or other molecules. The characterization of these conformational changes and their relation to protein function is a central goal of structural biology. Unfortunately, most conventional methods to obtain structural information do not provide information on protein dynamics. Therefore, mass spectrometry-based approaches, such as limited proteolysis, hydrogen−deuterium exchange, and stable-isotope labeling, are frequently used to characterize protein conformation and dynamics, yet the interpretation of these data can be cumbersome and time consuming. Here, we present PepShell, a tool that allows interactive data analysis of mass spectrometry-based conformational proteomics studies by visualization of the identified peptides both at the sequence and structure levels. Moreover, PepShell allows the comparison of experiments under different conditions, including different proteolysis times or binding of the protein to different substrates or inhibitors.

Limited Proteolysis Combined with Stable Isotope Labeling Reveals Conformational Changes in Protein (Pseudo)kinases upon Binding Small Molecules

“…The PDB entry 3OG7 was used for KD visualization. Images were generated in Pymol (The PyMOL Molecular Graphics System, version 1.5.0.4, Schrödinger) and by using Pepshell . (B) Histogram depicting the frequency of the pairwise correlation coefficients obtained by MD simulations for the B-RafWT complexed with ADP, ATP, AMP-PNP, and vemurafenib.…”

“…Cartoon representation of the RBD and KD of B-Raf WT and B-Raf V600E to illustrate the changes in abundance of peptides obtained in the limited proteolysis experiments of B-Raf WT complexed with small molecules (incubation for 4 h at 30 °C with 10 nmol ADP, ATP, or AMP-PNP) or its inhibitors (1 nmol vemurafenib or sorafenib) by using Pepshell. 83 The PDB entries 3NY5 and 3OG7 84 were used to visualize the RBD and the KD, respectively. Only the detected peptides that showed a statistically significant difference versus the control samples according to Student's t test (p < 0.…”

“…Figure 2. Conformational changes in B-Raf kinases induced by small-molecule binding by limited proteolysis coupled to stable isotope labeling MS.Cartoon representation of the RBD and KD of B-Raf WT and B-Raf V600E to illustrate the changes in abundance of peptides obtained in the limited proteolysis experiments of B-Raf WT complexed with small molecules (incubation for 4 h at 30 °C with 10 nmol ADP, ATP, or AMP-PNP) or its inhibitors (1 nmol vemurafenib or sorafenib) by using Pepshell 83. The PDB entries 3NY5 and 3OG784 were used to visualize the RBD and the KD, respectively.…”