Multivalent recognition of ferritin by full-length NCOA4 enables robust ferritinophagy.

Ferritinophagy is a central pathway in cellular iron homeostasis, yet the molecular basis by which the selective autophagy receptor NCOA4 recognizes and engages ferritin remains poorly defined, largely due to the long-standing inability of working with a soluble form of the full-length human NCOA4 (NCOA4FL). Here, we present an integrated biochemical and biophysical analysis of NCOA4FL and its interaction with human ferritin, complemented by structure-guided modeling. We show that NCOA4FL is predominantly intrinsically disordered and exists in a dynamic monomer-dimer equilibrium in solution, yet forms a stable, nanomolar-affinity complex with ferritin. Using crosslinking mass spectrometry, together with integrative modeling informed by low-resolution structural context, we demonstrate that NCOA4FL engages ferritin through a multivalent, distributed interface that is not confined to a short motif, in contrast to prior fragment-based models. Multiple interaction hotspots are identified on ferritin helices B and D, while NCOA4FL employs both central and terminal regions to wrap around the ferritin nanocage, enabling avidity-driven recognition. We further show that NCOA4FL coordinates a redox-sensitive [4Fe-4S] cluster, introducing a chemically defined feature that may contribute to its conformational plasticity and regulation. Together, these findings establish NCOA4FL as a flexible, multivalent ferritin receptor and define the recognition logic by which ferritin assemblies are selectively recognized and targeted for lysosomal degradation during ferritinophagy.