|Name||Mr. Samiol Azam|
|Organization||Florida International University|
Lead tightly associates with neuronal calcium sensor (NCS) protein DREAM and promotes structural changes analogous to calcium bound DREAM.
Samiol Azam, Jaroslava Miksovska
Florida International University, Miami, Fl, USA
Downstream regulatory element antagonist modulator (DREAM), is a 29kDa Ca2+ binding protein that controls several neurological processes including gene expression, apoptosis, and modulation of Kv4 voltage channels. Here we demonstrate that Pb2+ binds to EF-hands in DREAM with and equilibrium affinity higher that that determined for Ca2+. Pb2+ association triggers changes in the secondary and tertiary structure of the protein that are analogous to those observed in Ca2+ bound DREAM based on Trp 169 emission data and CD spectra. Namely, Pb2+ binding to DREAM leads to decrease of Trp 169 emission intensity and decrease in CD signal at 220nm. The hydrophobic cavity in the C-terminal domain of DREAM is solvent exposed in the presence of Pb2+ as determined using a hydrophobic probe 1, 8-ANS. The Kd values for 1,8-ANS binding to Ca2+ and Pb2+ bound DREAM were found to be similar; 73±10 μM and 103 ± 10 µM, respectively. Pb2+ binding to DREAM also modulates DREAM interactions with intracellular partners. For example, titrations of presenilin-1 with DREAM in the presence of Ca2+ or Pb2+ showed that Pb2+ bound DREAM has a similar affinity for presenilin-1 (Kd= 2.44±0.19 µM) as Ca2+ bound DREAM(Kd= 6.23 ± 1.60 µM). The impact of Pb2+ association on DREAM conformational dynamics was proved in time-resolved fluorescence studies and thermodynamics parameters for Pb2+ association to EF-hands were obtained using isothermal titration calorimetry (ITC). Those results indicate that DREAM and likely other neuronal calcium sensors may contribute to lead-induced toxicity.