Nontargeted Identification of Reactive Metabolite Protein Adducts
Michael G. Leeming
William A. Donald
Richard A. J. O’Hair
10.1021/acs.analchem.6b04604.s002
https://acs.figshare.com/articles/dataset/Nontargeted_Identification_of_Reactive_Metabolite_Protein_Adducts/5015822
Metabolic bioactivation
of many different chemicals results in
the formation of highly reactive compounds (chemically reactive metabolites,
CRMs) that can lead to toxicity via binding to macromolecular targets
(e.g., proteins or DNA). There is a need to develop robust, rapid,
and nontargeted analytical techniques to determine the identity of
the protein targets of CRMs and their sites of modification. Here,
we introduce a nontargeted methodology capable of determining both
the identity of a CRM formed from an administered compound as well
as the protein targets modified by the reactive metabolite in a single
experiment without prior information. Acetaminophen (<i>N</i>-acetyl-<i>p</i>-aminophenol, APAP) and <sup>13</sup>C<sub>6</sub>-APAP were incubated with rat liver microsomes, which are
known to bioactivate APAP to the reactive metabolite <i>N</i>-acetyl-<i>p</i>-benzoquinone imine (NAPQI). Global tryptic
digestion followed by liquid chromatographic/mass spectrometric (LC/MS)
analysis was used to locate “twin” ion peaks of peptides
adducted by NAPQI and for shotgun proteomics via tandem mass spectrometry
(MS/MS). By the development of blended data analytics software called
Xenophile, the identity of the amino acid residue that was adducted
can be established, which eliminates the need for specific parametrization
of protein database search algorithms. This combination of experimental
design and data analysis software allows the identity of a CRM, the
protein target, and the amino acid residues that are modified to be
rapidly established directly from experimental data. Xenophile is
freely available from https://github.com/mgleeming/Xenophile.
2017-05-08 00:00:00
compound
tandem mass spectrometry
data analytics software
Global tryptic digestion
adducted
acid
CRM
residue
data analysis software
NAPQI
Xenophile
MS
rat liver microsomes
13 C 6
reactive metabolite N
protein targets
APAP
DNA
Reactive Metabolite Protein Adducts Metabolic bioactivation
LC
acetyl
protein database search algorithms
identity
nontargeted