The applicant proposes a 2-AIM strategy to explore molecular mechanisms of SERCA calcium pump regulation by its major binding partner PLB.
AIM1 describes the use of fluorescence resonance energy transfer (FRET) to quantify the AFFINITY of PLB binding interactions and the STRUCTURE of the resulting protein complexes. The proposed experiments directly measure the effect of phosphorylation and mutation of PLB on its regulatory interactions. Because the fluorescent probes are genetically encoded, observations of these membrane protein interactions are being made in living cells for the first time.
AIM2 proposes to measure the PROTEIN BINDING KINETICS of the membrane protein complexes important for cardiac Ca2+ regulation. The rate of formation and dissolution of these membrane protein complexes is an important determinant of their function, but these rates are inaccessible to classical protein-protein binding kinetics methods. The applicant has invented a new optical method that can measure membrane protein subunit exchange dynamics in living cells.

Public Health Relevance

The proteins PLB and SERCA are of clinical significance because of their central role in cardiac function and disease. Disorders of these proteins are associated with heart failure, and loss or mutation of PLB in humans results in the disease """"""""dilated cardiomyopathy"""""""". Thus, PLB is a regarded as a high value therapeutic target in the treatment of heart failure, a leading cause of death in the United States.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL092321-05
Application #
8300136
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Krull, Holly
Project Start
2008-07-01
Project End
2013-06-30
Budget Start
2012-07-01
Budget End
2013-06-30
Support Year
5
Fiscal Year
2012
Total Cost
$330,784
Indirect Cost
$108,034
Name
Loyola University Chicago
Department
Physiology
Type
Schools of Medicine
DUNS #
791277940
City
Maywood
State
IL
Country
United States
Zip Code
60153
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Xu, Li; Pallikkuth, Sandeep; Hou, Zhanjia et al. (2011) Dysferlin forms a dimer mediated by the C2 domains and the transmembrane domain in vitro and in living cells. PLoS One 6:e27884
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