Phospholamban and its phosphorylated form interact differently with lipid bilayers: a 31P, 2H, and 13C solid-state NMR spectroscopic study.
Phospholamban (PLB) is a 52-amino acid integral membrane protein that helps to regulate the flow of Ca(2+) ions in cardiac muscle cells. Recent structural studies on the PLB pentamer and the functionally active monomer (AFA-PLB) debate whether its cytoplasmic domain, in either the phosphorylated or dephosphorylated states, is alpha-helical in structure as well as whether it associates with the lipid head groups (Oxenoid, K. (2005) Proc. Natl. Acad. Sci. U.S.A. 102, 10870-10875; Karim, C. B. (2004) Proc. Natl. Acad. Sci. U.S.A. 101, 14437-14442; Andronesi, C.A. (2005) J. Am. Chem. Soc. 127, 12965-12974; Li, J. (2003) Biochemistry 42, 10674-10682; Metcalfe, E. E. (2005) Biochemistry 44, 4386-4396: Clayton, J. C. (2005) Biochemistry 44, 17016-17026). Comparing the secondary structure of the PLB pentamer and its phosphorylated form (P-PLB) as well as their interaction with the lipid bilayer is crucial in order to understand its regulatory function. Therefore, in this study, the full-length wild-type (WT) PLB and P-PLB were incorporated into 1-palmitoyl-2-oleoyl-sn-glycero-phosphocholine (POPC) phospholipid bilayers and studied utilizing solid-state NMR spectroscopy. The analysis of the (2)H and (31)P solid-state NMR data of PLB and P-PLB in POPC multilamellar vesicles (MLVs) indicates that a direct interaction takes place between both proteins and the phospholipid head groups. However, the interaction of P-PLB with POPC bilayers was less significant compared that with PLB. Moreover, the secondary structure using (13)C=O site-specific isotopically labeled Ala15-PLB and Ala15-P-PLB in POPC bilayers suggests that this residue, located in the cytoplasmic domain, is a part of an alpha-helical structure for both PLB and P-PLB.
Abu-Baker
Shadi
S
Department of Chemistry and Biochemistry, Miami University, Oxford, Ohio 45056, USA.
Lorigan
Gary A
GA
D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002247:Carbon Isotopes; D003903:Deuterium; D008051:Lipid Bilayers; D019906:Nuclear Magnetic Resonance, Biomolecular; D010759:Phosphorus Isotopes; D010766:Phosphorylation; D011485:Protein Binding; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D002247:Carbon Isotopes; D003903:Deuterium; D008051:Lipid Bilayers; D010759:Phosphorus Isotopes; D000099211:Phospholamban