Cardiac sarcalumenin: Phosphorylation, comparison with the skeletal musclesarcalumenin and modulation of ryanodine receptor

Cardiac sarcoplasmic reticulum (SR) contains an endogenous phosphorylation system that under specific conditions phosphorylates two proteins with appa rent molecular masses of 150 and 130 kDa. The conditions for their phosphor ylation are as for the skeletal muscle sarcalumenin and the histidine-rich Ca2+ binding protein (HCP) with respect to: (i) Ca2+ and high concentration s of NaF are required; (ii) phosphorylation is obtained with no added Mg2and shows a similar time course and ATP concentration dependence; (iii) inh ibition by similar concentrations of La3+; (iv) phosphorylation is obtained with [gamma-P-32]GTP; (v) ryanodine binding is inhibited parallel to the p hosphorylation of the two proteins. The endogenous kinase is identified as casein kinase II (CK II) based on its ability to use GTP as effectively as ATP, and its inhibition by La3+. The association of CK II with the cardiac SR, even after EGTA extraction at alkaline pH, is demonstrated using antibo dies against CK II. The cardiac 130 kDa protein is identified as sarcalumen in based on its partial amino acid sequence and its blue staining with Stai ns-All. Cardiac sarcalumenin is different from the skeletal muscle protein based on electrophoretic mobilities, immunological analysis, peptide and ph osphopeptide maps, as well as amino acid sequencing. Preincubation of SR wi th NaF and ATP, but not with NaF and AMP-PNP caused strong inhibition of ry anodine binding. This is due to decrease in Ca2+- and ryanodine-binding aff inities of the ryanodine receptor (RyR) by about 6.6 and 18-fold, respectiv ely. These results suggest that cardiac sarcalumenin is an isoform of the s keletal muscle protein. An endogenous CK II can phosphorylate sarcalumenin, and in parallel to its phosphorylation the properties of the ryanodine rec eptor are modified.