Effects of Ni(II) and Cu(II) on DNA interaction with the N-terminal sequence of human protamine P2: enhancement of binding and mediation of oxidativeDNA strand scission and base damage

Epidemiological evidence suggests that certain paternal exposures to metals may increase the risk of cancer in the progeny. This effect may be associa ted with promutagenic damage to the sperm DNA, The latter is packed with pr otamines which might sequester carcinogenic metals and moderate the damage. Human protamine P2 has an amino acid motif at its N-terminus that can serv e as a heavy metal trap, especially for Ni(II) and Cu(II), We have synthesi zed a pentadecapeptide modeling this motif, Arg-Thr-His-Gly-Gln-Ser-His-Tyr -Arg-Arg-His-Cys-Ser-Arg-amide (HP2(1-15)) and described its complexes with Ni(II) and Cu(II), including their capacity to mediate oxidative DNA degra dation [Bal et al, (1997) Chern. Res. Toxicol,, 10, 906-914 and 915-921], I n the present study, effects of HP21-15 On NI(II)- and Cu(II)-mediated DNA oxidation by H2O2 at pH 7.4 were investigated in more detail using the circ ular plasmid pUC19 DNA as a target, and the single/double-strand breaks and production of oxidized DNA bases, as end points. Ni(II) alone was found to promote oxidative DNA strand scission (mostly single strand breaks) and ba se damage, while Cu(LI) alone produced the same effects, but to a much grea ter extent. Both metals were relatively more damaging to the pyrimidine bas es than to purine bases. HP2(1-15) fended to increase the Ni(H)/H2O2-induce d DNA breakage. In sharp contrast, the destruction of DNA strands by Cu(II) /H2O2 was almost completely prevented by HP2(1-15). The effect of HP21-15 O n the oxidative DNA base damage varied from a limited enhancement (5-hydrox yhydantoin and thymine glycol) to slight suppression (5-hydroxycytosine, 5- hydroxyuracil, 8-oxoguanine, 8-oxoadenine, 2-hydroxyadenine, fapyguanine an d fapyadenine) toward Ni(II)/H2O2. HP2(1-15) strongly suppressed the oxidat ive activity of Cu(II)/H2O2 in regard to all bases in DNA, Consistently wit h the above, the electron spin resonance/spin trap measurements revealed gr eater and more persistent generation of OH and O-2(-).-like oxidants from H 2O2 by the Ni(II)-HP2(1-15) complex than by the Cu(II)-HP2(1-15) complex (n o O-2(-). was detected). Both complexes were also found to bind to DNA more strongly than HP2(1-15) alone. The results indicate that protamine P2 is c apable of binding Ni(II) and Cu(II) and, in this way, attenuating the media tion of oxidative DNA damage by Cu(II), but not Ni(II), The effects found m ay be mechanistically involved in the reproductive toxicity and carcinogeni city of metals.