Treatment with divalent metal ions such as cobalt (Co2+) or nickel (Ni2+) result in the stabilization of hypoxia-inducible factor-1alpha (HIF1alpha). Recently, HIF1alpha was shown to be ubiquitinated by an E3-ligase complex and be subsequently targeted for proteasomal degradation. In this study, we demonstrated that Co2+ and Ni2+ specifically bind to cullin-2. Mutant analysis revealed that cullin-2 possesses at least three sites for the binding. Furthermore, fluorescence spectroscopy revealed that only Co2+ and Ni2+ have the binding activity to cullin-2, but other metal ions, including Cu2+, Ca2+, Mg2+, Mn2+, and Zn2+, did not. Finally, we found that Co2+ and Ni2+ do not bind to any components of the E3-ligase other than cullin-2, suggesting that cullin-2 is a key target of Co2+ and Ni2+. Interestingly, Co2+ did not affect the complex formation of the ligase, suggesting that the metal binding to cullin-2 affects the function, but not the assembly of the E3-ligase. (C) 2002 Elsevier Science.