EPR Accessibility Measurements of the SERCA-PLB Complex using Site-Directed Spin Labeling

Abstract

We investigate the structure formed byof phospholamban (PLB) bound to the sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA) using site-directed spin labeling. SERCA is a 994 residue transmembrane protein that helps control Ca2+ concentration in the SR relative to the cytoplasm in order to facilitate muscle relaxation. PLB is a 52 residue regulatory muscle protein found in cardiac muscle which, upon binding, has been shown to inhibit SERCA activity. However, phosphorylation of PLB at Ser-16 partially relieves this inhibition while still maintaining a combined structureconserving the complex. The structural mechanism behind this relief of inhibition is largely unknown. Using EPR accessibility measurements, we have shown that phosphorylation of PLB at Ser-16 alters the binding interface between its transmembrane helix and the PLB binding groove on SERCA, likely breaking the interactions that lead to inhibition.