Chronic
parasitic infections are a major risk factor for
cancer development in many underdeveloped countries. Oxidative damage of
DNA may provide a mechanism linking these processes. Eosinophil recruitment is a hallmark of
parasitic infections and many forms of
cancer, and
eosinophil peroxidase (EPO), a secreted hemoprotein, plays a central role in
oxidant production by these cells. However, mechanisms through which EPO may facilitate
DNA oxidation have not been fully characterized. Here, we show that EPO effectively uses plasma levels of
bromide as a cosubstrate to brominate bases in
nucleotides and
double-stranded DNA, forming several stable novel brominated adducts. Products were characterized by HPLC with on-line UV spectroscopy and electrospray ionization tandem mass spectrometry (LC/ESI/MS/MS). Ring assignments for brominated
purine bases as their 8-bromo adducts were identified by NMR spectroscopy. Using stable
isotope dilution LC/ESI/MS/MS, we show that while
guanine is the preferred
purine targeted for bromination as a free nucleobase,
8-bromoadenine is the major
purine oxidation product generated following exposure of
double-stranded DNA to either
HOBr or the EPO/H(2)O(2)/Br(-) system. Bromination of nucleobases was inhibited by scavengers of hypohalous
acids such as the
thioether methionine, but not by a large molar excess of primary
amines. Subsequently, N-monobromoamines were demonstrated to be effective brominating agents for both free nucleobases and
adenine within intact
DNA. A rationale for selective modification of
adenine, but not
guanine, in
double-stranded DNA based upon stereochemical criteria is presented. Collectively, these results suggest that specific brominated
DNA bases may serve as novel markers for monitoring oxidative damage of
DNA and the
nucleotide pool by brominating
oxidants.