Interaction of divalent metal ions with human translocase of inner membrane of mitochondria Tim23

The preprotein translocase of the inner membrane of mitochondria (TIM23 complex) is the main entry gate for proteins of the matrix and the inner membrane. Tim23p, the core component of TIM23 complex, forms the import pore across the inner membrane and exerts a key function in the protein import. However, the interaction of divalent metal ions with Tim23p and the contribution in the interaction of presequence peptide with Tim23p are still unknown. Herein, we investigated the interaction of divalent metal ions with the intermembrane space domain of Tim23p (Tim23IMS) and the interaction of presequence peptides with Tim23IMS in presence of Ca2+ ion by fluorescence spectroscopy in vitro. The static fluorescence quenching indicates the existence of strong binding between divalent metal ions and Tim23IMS. The order of the binding strength is Ca2+, Mg2+, Cu2+, Mn2+, and Co2+ (from strong to weak). Moreover, the interaction of presequence peptides with Tim23IMS is weakened in presence of Ca2+ ion, which implicates that Ca2+ ion may play an important role in the protein import by TIM23 complex. •Tim23p is the central component of TIM23 complex.•Interaction of Tim23p with presequence is crucial for precursor proteins importing.•We first found the strong interactions of divalent metal ions with human Tim23IMS.•The interaction of presequence with Tim23IMS is weakened in presence of Ca2+ ions.•It implicated that Ca2+ may play an important role in protein import by TIM23 complex.