Dissecting the structural and functional roles of a putative metal entry site in encapsulated ferritins
Cecilia Piergentili, Jennifer Ross, Didi He, Kelly J. Gallagher, Will A. Stanley, Laurène Adam, C. Logan Mackay, Arnaud Baslé, Kevin J. Waldron, David J. Clarke, Jon Marles‐Wright
Abstract
encapsulated ferritin at the interface between the two-helix subunits and proximal to the ferroxidase center. Here we present a comprehensive structural and functional study to investigate the functional relevance of this putative iron-entry site by means of enzymatic assays, MS, and X-ray crystallography. We show that catalysis occurs in the ferroxidase center and suggest a dual role for the secondary site, which both serves to attract metal ions to the ferroxidase center and acts as a flow-restricting valve to limit the activity of the ferroxidase center. Moreover, confinement of encapsulated ferritins within the encapsulin nanocage, although enhancing the ability of the encapsulated ferritin to undergo catalysis, does not influence the function of the secondary site. Our study demonstrates a novel molecular mechanism by which substrate flux to the ferroxidase center is controlled, potentially to ensure that iron oxidation is productively coupled to mineralization.