Structural constraints on the transmembrane and juxtamembrane regions of the phospholamban pentamer in membrane bilayers: Gln29 and Leu52.
The Ca2+-ATPase of cardiac muscle cells transports Ca2+ ions against a concentration gradient into the sarcoplasmic reticulum and is regulated by phospholamban, a 52-residue integral membrane protein. It is known that phospholamban inhibits the Ca2+ pump during muscle contraction and that inhibition is removed by phosphorylation of the protein during muscle relaxation. Phospholamban forms a pentameric complex with a central pore. The solid-state magic angle spinning (MAS) NMR measurements presented here address the structure of the phospholamban pentamer in the region of Gln22-Gln29. Rotational echo double resonance (REDOR) NMR measurements show that the side chain amide groups of Gln29 are in close proximity, consistent with a hydrogen-bonded network within the central pore. 13C MAS NMR measurements are also presented on phospholamban that is 1-13C-labeled at Leu52, the last residue of the protein. pH titration of the C-terminal carboxyl group suggests that it forms a ring of negative charge on the lumenal side of the sarcoplasmic reticulum membrane. The structural constraints on the phospholamban pentamer described in this study are discussed in the context of a multifaceted mechanism for Ca2+ regulation that may involve phospholamban as both an inhibitor of the Ca2+ ATPase and as an ion channel.
Liu
Wei
W
Department of Biochemistry and Cell Biology, Center for Structural Biology, Stony Brook University, Stony Brook, NY 11794-5115, USA.
Fei
Jeffrey Z
JZ
Kawakami
Toru
T
Smith
Steven O
SO
D000595:Amino Acid Sequence; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D005973:Glutamine / Q000737:chemistry* / Q000235:genetics; D006860:Hydrogen Bonding; D007930:Leucine / Q000737:chemistry* / Q000235:genetics; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D005973:Glutamine; D007930:Leucine; D008051:Lipid Bilayers; D008565:Membrane Proteins; D000099211:Phospholamban