Abstract
We report here the construction of phenanthroline (Phen) and terpyridine (Terpy)-based hybrid coordination motifs (HCMs), which were installed on the surface of the four-helical bundle hemeprotein cytochrome cb(562). The resulting constructs, termed HPhen1, HPhen2, HPhen3, and HTerpy1, feature HCMs that are composed of a histidine ligand and a Phen or Terpy functionality located two helix turns away, yielding stable tri- or tetradentate coordination platforms. Our characterization of the tridentate HCMs indicates that they accommodate many divalent metal ions (Co(2+), Ni(2+), Cu(2+), Zn(2+)) with nanomolar to femtomolar affinities, lead to significant stabilization of the alpha-helical protein scaffold through metal-mediated cross-linking, assert tight control over protein dimerization, and provide stable and high-affinity binding sites for substitution-inert metal probes. Our analyses suggest that such tridentate HCMs may be used modularly on any alpha-helical protein surface in a sequence-independent fashion.