Recent developments in instrumentation
and bioinformatics have
led to new quantitative mass spectrometry platforms including LC–MS/MS
with data-independent acquisition (DIA) and targeted analysis using
parallel reaction monitoring mass spectrometry (LC–PRM), which
provide alternatives to well-established methods, such as LC–MS/MS
with data-dependent acquisition (DDA) and targeted analysis using
multiple reaction monitoring mass spectrometry (LC–MRM). These
tools have been used to identify signaling perturbations in lung cancers
and other malignancies, supporting the development of effective kinase
inhibitors and, more recently, providing insights into therapeutic
resistance mechanisms and drug repurposing opportunities. However,
detection of kinases in biological matrices can be challenging; therefore,
activity-based protein profiling enrichment of ATP-utilizing proteins
was selected as a test case for exploring the limits of detection
of low-abundance analytes in complex biological samples. To examine
the impact of different MS acquisition platforms, quantification of
kinase ATP uptake following kinase inhibitor treatment was analyzed
by four different methods: LC–MS/MS with DDA and DIA, LC–MRM,
and LC–PRM. For discovery data sets, DIA increased the number
of identified kinases by 21% and reduced missingness when compared
with DDA. In this context, MRM and PRM were most effective at identifying
global kinome responses to inhibitor treatment, highlighting the value
of a priori target identification and manual evaluation of quantitative
proteomics data sets. We compare results for a selected set of desthiobiotinylated
peptides from PRM, MRM, and DIA and identify considerations for selecting
a quantification method and postprocessing steps that should be used
for each data acquisition strategy
