Hemopexin (Hpx) is a plasma glycoprotein with very high affinity for heme. It can mitigate heme-mediated oxidative stress and cardiovascular dysfunction and has promising therapeutic applications for hemolytic disorders. The uptake of heme-Hpx is accompanied by an increase in intracellular Cu levels, thus hinting at a dynamic interaction between Cu and Hpx. In vitro studies have indicated that Cu(II) can disrupt heme-Hpx complex formation. Consequently, it has been proposed that Cu(II) binds to Hpx under the acidic endosomal conditions, preventing heme re-binding. This project seeks to elucidate the interplay between Cu and heme binding to Hpx. To this end, X-ray Absorption Spectroscopy will be used for the first time to characterize heme- and Cu-binding sites in Hpx in different pH conditions to understand (i) how Cu(II) is bound at both pH, (ii) whether it can be released upon reduction by physiological agents and (iii) what is the mutual impact of Cu- and heme-binding.