Structure-function relationships in the Ca2+- binding and translocation domain of SERCA1:: physiological correlates in Brody disease

Alanine-scanning mutagenesis of all amino acids in transmembrane helices M4, M5, M6 and M8, which contain known Ca2+ binding residues in the Ca2+-ATPase of skeletal muscle sarcoplasmic reticulum, revealed patches of mutation-sensitivity in M4, M5 and M6, but not in M8. A six-residue motif, (E/D)GLPA(T/V), in M4 and M6 and its counterpart in M5 were highlighted by mutagenesis. Site-directed disulfide mapping of helices M4 and M6 demonstrated that these transmembrane helices associate as a right-handed coiled-coil. This structural information, combined with the earlier analysis of the association of each Ca2+ binding residue with either Ca2+ binding site I or site II, permitted the development of a "side-by-side" model for the two Ca2+ binding sites in the Ca2+-ATPase. In about half of Brody disease families, mutations create stop codons which delete all or part of the Ca2+ binding and translocation domain, resulting in loss of SERCA1 function and muscle disease.