Exploring the shifts in the energy landscape ofcalmodulin under different environmental conditions

Proteins are flexible structures, and their functions are closely tied to their ability to adoptmultiple conformations. Their dynamic nature enables them to shift between different shapesor states, which is crucial for interacting with other biomolecules or catalyzing reactions.This behavior of proteins is strongly influenced by environmental conditions, such astemperature, pH, and ionic strength, which can significantly alter their structural states andfunctions.In this thesis, we investigate how the conformational landscape of proteins shiftsunder changing environmental conditions. Our model system is calmodulin (CaM), acalcium-binding protein involved in numerous cellular processes. Known for its adaptability,CaM exhibits a wide array of conformations under various experimental conditions, makingit an ideal candidate for studying environmental effects on protein dynamics. Moreover, CaMserves as the sensing domain in genetically encoded fluorescent calcium biosensors,highlighting the importance of understanding its conformational changes for designingefficient and responsive sensors. The regions visited by CaM are well-represented by a pairof essential degrees of freedom (DoF) describing the relative positioning of its two lobes(cis/trans) and the compactness of the overall CaM structure. Since the time scale of classicalmolecular dynamics (MD) simulations do not allow for overcoming the high energy barriersthat might separate various minima, we resort to well-tempered metadynamics simulationsusing the essential DoF as collective variables (CVs). We explore four different conditionsrepresenting Ca2+ bound/unbound CaM at physiological/low ionic strength. We find that CaM acts like a juggler, adjusting the position of its minima between four stable regions (ciscompact,trans-compact, cis-open, trans-open). After identifying the locations of the minimavia metadynamics, we select representative structures from these minima and performequilibration runs. Our work unveils the structural basis of the observed minima and validatesthe effectiveness of our chosen CVs. Our results reveal that, although the energy surface ismuch shallower under low ionic strength (IS) conditions, conformers get stuck in artificialminima due to strong transient salt bridges. It appears that ions act as lubricants, helping torelease salt bridges that cause the conformers to become trapped. As a result, we find IS ofthe environment to be a major factor to be considered in protein design problems.