Revisiting the interaction of heme with hemopexin
Author:
Detzel Milena Sophie1, Schmalohr Benjamin Franz1, Steinbock Francèl1, Hopp Marie-Thérèse1, Ramoji Anuradha23, Paul George Ajay Abisheck1, Neugebauer Ute23, Imhof Diana1ORCID
Affiliation:
1. Pharmaceutical Biochemistry and Bioanalytics , Pharmaceutical Institute, University of Bonn , An der Immenburg 4 , D-53121 Bonn , Germany 2. Center for Sepsis Control and Care (CSCC) , Jena University Hospital , D-07747 Jena , Germany 3. Leibniz Institute of Photonic Technology , Albert-Einstein-Str. 9 , D-07745 Jena , Germany
Abstract
Abstract
In hemolytic disorders, erythrocyte lysis results in massive release of hemoglobin and, subsequently, toxic heme. Hemopexin is the major protective factor against heme toxicity in human blood and currently considered for therapeutic use. It has been widely accepted that hemopexin binds heme with extraordinarily high affinity of <1 pM in a 1:1 ratio. However, several lines of evidence point to a higher stoichiometry and lower affinity than determined 50 years ago. Here, we re-analyzed these data. SPR and UV/Vis spectroscopy were used to monitor the interaction of heme with the human protein. The heme-binding sites of hemopexin were characterized using hemopexin-derived peptide models and competitive displacement assays. We obtained a K
D
value of 0.32 ± 0.04 nM and the ratio for the interaction was determined to be 1:1 at low heme concentrations and at least 2:1 (heme:hemopexin) at high concentrations. We were able to identify two yet unknown potential heme-binding sites on hemopexin. Furthermore, molecular modelling with a newly created homology model of human hemopexin suggested a possible recruiting mechanism by which heme could consecutively bind several histidine residues on its way into the binding pocket. Our findings have direct implications for the potential administration of hemopexin in hemolytic disorders.
Publisher
Walter de Gruyter GmbH
Subject
Clinical Biochemistry,Molecular Biology,Biochemistry
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