PEP-19 is a small, intrinsically disordered protein (IDP) that regulates the binding kinetics of the protein calmodulin (CaM) to calcium. Calmodulin is a calcium signal messenger, responding to changes in intracellular calcium levels to regulate hundreds of different pathways. PEP-19’s role is crucial, with inappropriate expression leading to calcium signaling disruption that can contribute to neurodegenerative diseases, cardiac hypertrophy, and cancers. In the brain, its presence helps to protect against calcium overload, with increased concentrations found in areas spared from Alzheimer’s disease, and decreased concentrations found in Huntington’s disease. Despite playing such an essential role, little is known about the conformational ensemble of PEP-19 and how it affects calmodulin binding to its target proteins. Our goal is to study this conformational ensemble using fluorescence resonance energy transfer (FRET) fluorometry, labeling with the organic fluorophore IAEDANS. This allows us to measure the end-to-end distance of PEP-19 in order to look for changes in its compaction or expansion. By measuring FRET in the presence and absence of different crowding agents, denaturants, and osmolytes, we can observe the effect of the local environment on the PEP-19 conformational ensemble.

Investigating the conformational ensemble of calmodulin-binding protein PEP-19

ACS Spring 2022

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