1 research outputs found
Analysis of Covalent Modifications of Proteins by Oxidized Phospholipids Using a Novel Method of Peptide Enrichment
Free
radical-induced oxidation of phospholipids contributes significantly
to pathologies associated with inflammation and oxidative stress.
Detection of covalent interaction between oxidized phospholipids (oxPL)
and proteins by LC-MS/MS could provide valuable information about
the molecular mechanisms of oxPL effects. However, such studies are
very limited because of significant challenges in detection of the
comparatively low levels of oxPL–protein adducts in complex
biological systems. Current approaches have several limitations, most
important of which is the inability to detect protein modifications
by naturally occurring oxPL. We now report, for the first time, an
enrichment method that can be applied to the global analysis of protein
adducts with various naturally occurring oxPL in relevant biological
systems. This method exploits intrinsic properties of peptides modified
by oxPL, allowing highly efficient enrichment of oxPL-modified peptides
from biological samples. Very low levels of oxPL–protein adducts
(<2 ppm) were detected using this enrichment method in combination
with LC-MS/MS. We applied the method to several model systems, including
oxidation of high density lipoprotein (HDL) and interaction of human
platelets with a specific oxPL, and demonstrated its extremely high
efficiency and productivity. We report multiple new modifications
of apolipoproteins in HDL and proteins in human platelets