Pulse dipolar EPR spectroscopy reveals buffer modulated cooperativity of metal templated protein dimerization

Self-assembly of protein monomers directed by metal ion coordination constitutes a promising strategy for designing supramolecular architectures complicated by the non-covalent interaction between monomers. Herein, two pulse dipolar electron paramagnetic resonance spectroscopy (PDS) techniques, double electron-electron resonance (DEER) and relaxation-induced dipolar modulation enhancement (RIDME), were simultaneously employed for studying the CuII-templated dimerization behavior of a model protein (Streptococcus sp. Group G, Protein G B1 domain) in both phosphate and Tris-HCl buffers. A cooperative binding model could simultaneously fit all data and demonstrate cooperativity of protein dimerization across α-helical double-histidine motifs in presence of Cu(II) is strongly buffer modulated, representing a platform for highly tunable buffer-switchable templated dimerization. Hence, PDS enriches the family of techniques for monitoring binding processes, supporting the development of novel strategies for bioengineering structures and stable architectures assembled by an initial metal-templated dimerization.