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Ferritin is a 24-mer protein nanocage that stores iron and regulates intracellular iron homeostasis. The nuclear receptor coactivator-4 (NCOA4) binds specifically to ferritin H subunits and facilitates the autophagic trafficking of ferritin to the lysosome for degradation and iron release. Using isothermal titration calorimetry (ITC), we studied the thermodynamics of the interactions between ferritin and the soluble fragment of NCOA4 (residues 383–522), focusing on the effects of the recently identified Fe–S cluster bound to NCOA4, ferritin subunit composition, and ferritin-iron loading. Our findings show that in the presence of the Fe–S cluster, the binding is driven by a more favorable enthalpy change and a decrease in entropy change, indicating a key role for the Fe–S cluster in the structural organization and stability of the complex. The ferritin iron core further enhances this association, increasing binding enthalpy and stabilizing the NCOA4-ferritin complex. The ferritin subunit composition primarily affects binding stoichiometry of the reaction based on the number of H subunits in the ferritin H/L oligomer. Our results demonstrate that both the Fe–S cluster and the ferritin iron core significantly affect the binding thermodynamics of the NCOA4-ferritin interactions, suggesting regulatory roles for the Fe–S cluster and ferritin iron content in ferritinophagy.