The regulation of calcium levels across the membrane of the sarcoplasmic re
ticulum involves the complex interplay of several membrane proteins. Phosph
olamban is a 52 residue integral membrane protein that is involved in rever
sibly inhibiting the Ca2+ pump and regulating the flow of Ca ions across th
e sarcoplasmic reticulum membrane during muscle contraction and relaxation.
The structure of phospholamban is central to its regulatory role. Using ho
monuclear rotational resonance NMR methods, we show that the internuclear d
istances between [1-C-13]Leu7 and [3-C-13]Ala11 in the cytoplasmic region,
between [1-C-13]Pro21 and [3-C-13]Ala24 in the juxtamembrane region and bet
ween [1-C-13]Leu42 and [3-C-13]Cys46 in the transmembrane domain of phospho
lamban are consistent with alpha -helical secondary structure. Additional h
eteronuclear rotational-echo double-resonance NMR measurements confirm that
the secondary structure is helical in the region of Pro21 and that there a
re no large conformational changes upon phosphorylation. These results supp
ort the model of the phospholamban pentamer as a bundle of five long alpha
-helices. The long extended helices provide a mechanism by which the cytopl
asmic region of phospholamban interacts with residues in the cytoplasmic do
main of the Ca2+ pump. (C) 2001 Academic Press.