Defining the intramembrane binding mechanism of sarcolipin to calcium ATPase using solution NMR spectroscopy.
Sarcolipin (SLN) is an integral membrane protein that is expressed in both skeletal and cardiac muscle, where it inhibits SERCA (calcium ATPase) by lowering its apparent Ca2+ affinity in a manner similar to that of its homologue phospholamban (PLN). We use solution NMR to map the structural changes occurring within SLN upon interaction with the regulatory target, SERCA, co-reconstituting the two proteins in dodecylphosphocholine (DPC) detergent micelles, a system that preserves the native structure of SLN and the activity of SERCA, with the goal of comparing these interactions with those of the previously studied PLN-SERCA complex. Our analysis of the structural dynamics of SLN in DPC micelles shows this polypeptide to be partitioned into four subdomains: a short unstructured N terminus (residues 1-6), a short dynamic helix (residues 7-14), a more rigid helix (residues 15-26), and an unstructured C terminus (residues 27-31). Upon addition of SERCA, the different domains behave according to their dynamics, molding onto the surface of the enzyme. Remarkably, each domain of SLN behaves in a manner similar to that of the corresponding domains in PLN, supporting the hypothesis that both SLN and PLN bind SERCA in the same groove and with similar mechanisms.
Buffy
Jarrod J
JJ
Department of Chemistry, University of Minnesota, Minneapolis, MN 55455, USA.
Buck-Koehntop
Bethany A
BA
Porcelli
Fernando
F
Traaseth
Nathaniel J
NJ
Thomas
David D
DD
Veglia
Gianluigi
G
D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D009124:Muscle Proteins / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011510:Proteolipids / Q000378:metabolism*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
D008823:Micelles; D009124:Muscle Proteins; D011510:Proteolipids; C077258:sarcolipin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium