Structural heterogeneity of iron–sulfur cluster scaffold protein IscU: Metamorphic or partially folded?

The scaffold protein IscU plays a central role in iron-sulfur (Fe-S) cluster assembly and transfer. Despite extensive structural and biochemical research, the conformational nature of IscU remains unclear. IscU has been proposed as a metamorphic protein that interconverts between structured (S) and disordered (D) states. However, its conformational heterogeneity has often been interpreted as a consequence of intrinsic marginal stability, with the D-state described as an unfolded and functionally irrelevant state. These contrasting views highlight the fundamental challenge in defining the structural identity of IscU. In this review, we provide a comprehensive overview of the structural heterogeneity of IscU and examine how intrinsic and extrinsic factors shape the conformational equilibrium between the S- and D-states. Furthermore, we consider how these conflicting interpretations could be reconciled within an ensemble-based framework. In addition, we highlight promising approaches for characterizing the structurally elusive D-state, including high-pressure nuclear magnetic resonance, single-molecule Förster resonance energy transfer, native ion mobility mass spectrometry, and artificial intelligence-based structural analysis, all of which provide new opportunities for probing low-populated and transient conformational states. Collectively, these perspectives suggest that the functional behavior of IscU is closely linked to its conformational heterogeneity and that a conformational landscape view may provide a more integrated understanding of its role in Fe-S cluster biosynthesis.