|Organization or Institution||Florida International University|
Role of F235A residue in neuronal calcium sensor DREAM
Maria Santiago, Maria Daniel Santiago, Jaroslava Miksovska
Florida International University. Department and Chemistry and Biochemistry
Neuronal calcium sensors proteins are in charge of regulating a variety of physiological processes in the brain and they are ultimately linked to several pathological conditions such as Parkinson and Alzheimer disease. Downstream regulatory element antagonist modulator (DREAM) is part of the neuronal calcium sensor family. DREAM is an EF-hand protein which is highly expressed in various areas in the central nervous system such as the hippocampus and the neural cortex. It is known to have a crucial role in regulating the kinetics of potassium channels, gene expression, calcium homeostasis and enzymatic activity of presenilin in the cytoplasm. This diverse functionality of DREAM is due to its interaction with multiple intracellular partners. Previous studies have demonstrated that DREAM affinity for DNA and for potassium voltage channel site 2 are modulated by the presence of Ca2+ suggesting that Ca2+ function as an allosteric modulate. The precise mechanism of how Ca2+ regulates DREAM interactions with effector partners is unclear as the Ca2+ binding sites are located in the C- termini and the DNA and Kv channel site 2 binding sites are found at the N- terminal domain. Based on the molecular dynamic data we propose that a network of hydrophobic residues connects the C- and N- terminal domain and Phe235 is a key residue in the interdomain communication. With this in mind, we have prepared a DREAM construct with Phe235 replaced by Ala and characterized its photophysical properties as well as its affinity for intracellular partners. Our results show that Ca2+ association to Phe235Ala DREAM leads to distinct conformational changes compare to the WT protein. More importantly, the affinity of Phe235Ala DREAM for site 2 is significantly diminished compare to the WT confirming the role of Phe 235 in the calcium signal transduction mechanism.