Adductomics: characterizing exposures to reactive electrophiles

Abstract

To understand environmental causes of disease, unbiased methods are needed to characterize the human exposome, which represents all toxicants to which people are exposed from both exogenous and endogenous sources. Because they directly modify DNA and important proteins, reactive electrophiles are probably the most important constituents of the exposome. Exposures to reactive electrophiles can be characterized by measuring adducts from reactions between circulating electrophiles and blood nucleophiles. We define an 'adductome' as the totality of such adducts with a given nucleophilic target. Because of their greater abundance and residence times in human blood, adducts of hemoglobin (Hb) and human serum albumin (HSA) are preferable to those of DNA and glutathione for characterizing adductomes. In fact, the nucleophilic hotspot represented by the only free sulfhydryl group in HSA (HSA-Cys(34)) offers particular advantages for adductomic experiments. Although targeted adducts of HSA-Cys(34) have been monitored for decades, an unbiased method has only recently been reported for visualizing the HSA-Cys(34) 'subadductome'. The method relies upon a novel mass spectrometry application, termed fixed-step selected reaction monitoring (FS-SRM), to profile Cys(34) adducts in tryptic digests of HSA. Here, we selectively review the literature regarding the potential of adductomics to partially elucidate the human exposome, with particular attention to the HSA-Cys(34) subadductome.

Fig. 1

Levels of HSA adducts of benzene oxide (BO-Alb) and 1,4-benzoquinone (1,4-BQ-Alb) versus exposure to benzene for 439 Chinese workers (Lin et al., 2007; Rappaport et al., 2002; Yeowell-O’Connell et al., 1998). Relative adduct levels (RALs) represent ratios of individual adduct levels to median levels in concurrent controls.

Fig. 2

Humans are exposed to reactive and potentially toxic electrophiles from both exogenous and endogenous sources. The HSA molecule contains a nucleophilic hotspot (HSA-Cys³⁴) that forms adducts with electrophiles in the blood and can be used to characterize human exposures to reactive electrophiles (or their precursors) during the month prior to blood collection.

Fig. 3

Scheme for profiling HSA-Cys³⁴ adducts (Li et al., 2011). Thiol-affinity resins are used to remove mercaptalbumin (i.e. HSA containing free Cys³⁴). Enriched HSA-Cys³⁴ adducts are purified by high-performance liquid chromatography (HPLC), detected as modified T3 peptides by fixed-step selected reaction monitoring (FS-SRM), and displayed as a profile of adduct concentration vs. added mass.

Fig. 4

HSA-Cys³⁴ adducts detected in archived HSA from subjects stratified by race, gender and smoking status (Li et al., 2011). Each HSA sample was pooled from 5 subjects stratified by race (B = black, W = white), gender (F = female, M = male, and smoking status (N = nonsmoker, S = smoker). Adduct-mass hits in each HSA sample are shown with estimated concentrations expressed as pmol/mg of HSA (represented by colors). Each adduct mass represents the midpoint of a mass bin ±2.3 Da.