The role of ATP, ADP and divalent cations in the formation of binary and ternary complexes of actin, cofilin and DNase I Actin is the major cytoskeletal protein of virtually all eukaryotic cells. Actin assembly/ disassembly is involved in a variety of cellular processes and actin-binding proteins are essential in regulation of the pool of actin monomers. Cofilin and DNase I are actinbinding proteins, which form both binary (actin-DNase 1, cofilin-actin) and ternary (cofilin-actin-DNase I) complexes with actin. Here we use native gel electrophoresis to examine the roles of ATP, ADP, Ca 2+ and Mg 2+ in the formation of these complexes as well as on the ability of actin to self-assemble. Conditions which favour actin polymerisation are: ATP (no Me 2+ ) ³ ADP (no Me 2+ ) > ADP-Ca 2+ = ADP-Mg 2+ > ATP-Mg 2+ > ATP-Ca 2+ . Preferential conditions for the formation of the binary actin-cofilin complex are: ADP-Mg 2+ ³ ADP-Ca 2+ > > ATP-Ca 2+ & ATP-Mg 2+ & ADP-No Me 2+ & ATP-No Me 2+ . Actin forms a very tight complex with DNase I in the order: ATP-Ca 2+ ³ ATP-Mg 2+ & ADP-Mg 2+ & ADP-Ca 2+ ³ ADP-(no Me 2+ ) > ATP-(no Me 2+ ). Effectively, the complex does not form in the presence of ATP and the absence of free Me 2+ . Finally, the conditions which favour the formation of a ternary complex of cofilin-actin-DNase I resemble the actin-DNase I, namely: ATP-Ca 2+ & ADP-Ca 2+ & ADP-Mg 2+ & ATP-Mg 2+ & ADP (no Me 2+ ) > ATP-(no Me 2+ ).