posted on 2021-10-04, 18:44authored byHe Zhu, Scott B. Ficarro, William M. Alexander, Laura E. Fleming, Guillaume Adelmant, Tinghu Zhang, Matthew Willetts, Jens Decker, Sven Brehmer, Michael Krause, Michael P. East, Nathanael S. Gray, Gary L. Johnson, Gary Kruppa, Jarrod A. Marto
Parallel
reaction monitoring (PRM) has emerged as a popular approach
for targeted protein quantification. With high ion utilization efficiency
and first-in-class acquisition speed, the timsTOF Pro provides a powerful
platform for PRM analysis. However, sporadic chromatographic drift
in peptide retention time represents a fundamental limitation for
the reproducible multiplexing of targets across PRM acquisitions.
Here, we present PRM-LIVE, an extensible, Python-based acquisition
engine for the timsTOF Pro, which dynamically adjusts detection windows
for reproducible target scheduling. In this initial implementation,
we used iRT peptides as retention time standards and demonstrated
reproducible detection and quantification of 1857 tryptic peptides
from the cell lysate in a 60 min PRM-LIVE acquisition. As an application
in functional proteomics, we use PRM-LIVE in an activity-based protein
profiling platform to assess binding selectivity of small-molecule
inhibitors against 220 endogenous human kinases.