While copper is an essential redox cofactor, excess copper, particularly Cu(I), is highly toxic and therefore requires precise control of metal speciation in the bacterial periplasm. CopI is a cupredoxin-like protein implicated in copper tolerance under microaerobic conditions and contains multiple histidine- and methionine-rich regions, yet its metal-binding properties remain poorly understood. Here, we deconstructRubrivivax gelatinosusCopI into its constituent metal-binding modules and provide the first quantitative, domain-resolved analysis of their Cu(II) coordination chemistry. Two peptide domains, an N-terminal His-rich region and an internal His/Met-rich motif, form mononuclear 1:1 Cu(II) complexes but exhibit strikingly different binding modes. The His-rich domain displays higher Cu(II) affinity and a rigid, predominantly nitrogen-based coordination environment, whereas the His/Met-rich motif binds Cu(II) more weakly and adopts a heterogeneous, N/S/O-donor coordination sphere indicative of structural flexibility. Neither domain forms stable secondary structure upon metal binding, indicating that intrinsic disorder is a functional feature rather than a structural deficiency. Comparative Zn(II) studies reveal substantially weaker binding, underscoring copper selectivity. Collectively, these results identify CopI as a chemically modular copper buffer and establish a coordination-chemical framework for understanding copper handling by periplasmic cupredoxin-like proteins.
Sobol et al. (Fri,) studied this question.