Electrophilic DNA adducts produced following oxidative stress can form DNA-protein cross-links (DPCs), dramatically altering genomic maintenance pathways. Complete characterization of DPCs has been hindered, in part, because of a lack of comprehensive techniques for their analysis. We have, therefore, established a proteomics approach to investigate sites of cross-link formation using N(6)-(3-oxo-1-propenyl)-2'-deoxyadenosine (OPdA), an electrophilic DNA adduct produced from oxidative stress. OPdA was reacted with albumin and reduced with NaBH4 to stabilize DPCs. Using LC-MS/MS proteomics techniques, high-resolution peptide sequence data were obtained; however, using a database searching strategy, adducted peptides were only identified in samples subjected to chemical depurination. This strategy revealed multiple oxopropenyl adenine-lysine adducts and oxopropenyl-lysine adducts with the most reactive lysines identified to be Lys256 and Lys548. Manual interrogation of the mass spectral data provided evidence of OPdA deoxynucleoside conjugates to lysines and cross-links that underwent facile collision-induced dissociation to release an unmodified peptide without subsequent fragmentation. These fragmentations precluded adduct detection and peptide sequencing using database searching methods. Thus, comprehensive analysis of DPCs requires chemical depurination of DNA-protein reaction mixtures followed by a combination of database-dependent and manual interrogation of LC-MS/MS data using higher-energy collision-induced dissociation. In the present case, this approach revealed that OPdA selectively modifies surface lysine residues and produces nucleoside-protein cross-links and oxopropenyl lysine.