Spectroscopic and Theoretical Study of CuI Binding to His111 in the Human Prion Protein Fragment 106-115

The ability of the cellular prion protein (PrPC) to bind copper in vivo points to a physiological role for PrPC in copper transport. Six copper binding sites have been identified in the nonstructured N-terminal region of human PrPC. Among these sites, the His111 site is unique in that it contains a MKHM motif that would confer interesting Cu-I and Cu-II binding properties. We have evaluated Cu-I coordination to the PrP(106-115) fragment of the human PrP protein, using NMR and X-ray absorption spectroscopies and electronic structure calculations. We find that Met109 and Met112 play an important role in anchoring this metal ion. Cu-I coordination to His111 is pH-dependent: at pH >8, 2N1O1S species are formed with one Met ligand in the range of pH 5-8, both methionine (Met) residues bind to Cu-I, forming a 1N1O2S species, where N is from His111 and O is from a backbone carbonyl or a water molecule; at pH <5, only the two Met residues remain coordinated. Thus, even upon drastic changes in the Chemical environment, such as those occurring during endocytosis of PrPC (decreased pH and a reducing potential), the two Met residues in the MKHM motif enable PrPC to maintain the bound Cu-I ions, consistent with a copper transport function for this protein. We also find that the physiologically relevant Cu-I-1N1O2S species activates dioxygen via an inner-sphere mechanism, likely involving the formation of a copper(II) superoxide complex. In this process, the Met residues are partially Oxidized to sulfoxide; this ability to scavenge superoxide may play a role in the proposed antioxidant properties of PrPC. This study provides further insight into the Cu-I coordination properties of His111 in human PrPC and the molecular mechanism of oxygen activation by this site.