Molecular modeling of fluorescent SERCA biosensors

Molecular modeling and simulation are useful tools in structural biology, allowing the formulation of functional hypotheses and interpretation of spectroscopy experiments. Here, we describe a method to construct in silico models of a fluorescent fusion protein construct, where a cyan fluorescent protein (CFP) is linked to the actuator domain of the Sarco/Endoplasmic Reticulum Ca²⁺-ATPase (SERCA). This CFP-SERCA construct is a biosensor that can report on structural dynamics in the cytosolic headpiece of SERCA. Molecular modeling and FRET experiments allow us to generate new structural and mechanistic models that better describe the conformational landscape and regulation of SERCA. The methods described here can be applied to the creation of models for any fusion protein constructs and also describe the steps needed to simulate FRET results using molecular models.