Crowded environments tune the fold-switching in metamorphic proteins

Metamorphic proteins such as circadian clock protein KaiB and human chemokine XCL1 play vital roles in regulating biological processes, including gene expression, circadian clock and innate immune responses, and perform distinct functions in living cell by switching different structures in response to cellular environment stimuli. However, it is unclear how complex and crowded intracellular environments affect conformational rearrangement of metamorphic proteins. Here, the kinetics and thermodynamics of two well-characterized metamorphic proteins, circadian clock protein KaiB and human chemokine XCL1, were quantified in physiologically relevant environments by using NMR spectroscopy, indicating that crowded agents shift equilibrium towards the inactive form (ground-state KaiB and Ltn10-like state XCL1) without disturbing the corresponding structures, and crowded agents have predominantly impact on the exchange rate of XCL1 that switches folds on timescales of seconds, but have slightly impact on the exchange rate of KaiB that switches folds on timescales of hours. Our data shed light on how metamorphic proteins can respond immediately to the changed crowded intracellular conditions that induced by environmental cues and then execute different functions in living cell, and it also enhances our understanding of how environments enrich the sequence-structure-function paradigm.