High Affinity Calcium Binding

Calcium ions are bound with an identical high affinity of 5.106M by the purified ATPase, by the transport protein in the native membranes as well as by partially deli-pidated, reversibly inactivated membrane preparations 8, ll9 173). The amount of calcium which is bound with that high affinity corresponds to two sites per transport molecule. The observed affinity is in good agreement with the affinity derived from the dependence on ionized calcium of the activation of calcium uptake and ATP splitting as well as of the inhibition of calcium release and ATP synthesis18 u2,, 74 17s Since the latter experiments were performed under conditions which provide a constant internal free calcium concentration by the presence of oxalate or phosphate in the system, the reactions must have been activated or inhibited by the calcium ions 

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P/Q-type channels undergo a second calcium-dependent process whereby current activity increases following a strong membrane depolarization or a train of action potentials. This increase is only observed in the presence of external calcium, is insensitive to calcium buffers, and requires the high-affinity calcium binding sites on calmodulin (Chaudhuri et al. 2004 DeMaria et al. 2001). This process, termed calcium-dependent facilitation (CDF), is not observed with N-type or R-type channels and is Cav2.1 channel splice isoform dependent (Chaudhuri et al. 2004). The... 

Parvalbumin is a small (12 kDa), cytoplasmic, high-affinity, calcium-binding protein that has structural similarity to TnC and calmodulin. It is found especially in striated muscles in an amount proportional to their speed of relaxation. It acts as a calcium buffer, facilitating muscle relaxation by transferring calcium ions from TnC to the Ca-pump that sequesters calcium within the sarcoplasmic reticulum. Parvalbumin concentration in urine has been found to be increased with myotoxicity (Dare et al. 2002). 

Calcium transport depends on the occupance of high-affinity calcium-binding sites on the external surface of the membranes (cf. [2,21,44,83- 85]). There is general agreement that two high-affinity calcium-binding sites are involved and that the saturation of these sites and the activation of transport and ATP hydrolysis occur in the same concentration range. 

Linse, S., Forsen, S. 1995. Determinants that govern high-affinity calcium binding. Adv Second Messenger Phosphoprotein Res, 30 89- 15 1. 

As already mentioned, calcium is the most important physiological effector of RyR activity (10 1I 23). High-affinity calcium binding sites have been described in matrunalian RyR subtypes (24 25). In addition, calcium regulation of ryanodine binding and channel opening of insect RyR has been reported previously ( II 12 26). 

Calcium and chloride cofactors have been shown to be needed for maximum rates of oxygen evolution [1,2]. The removal of calcium ions from PS2 has been linked to the loss of 17kDa and 23kDa extrinsic polypeptides. These polypeptides have been suggested to protect a high affinity calcium binding site. A low pH citrate wash has been used to remove one calcium per PS2 without loss of the extrinsic polypeptides [3]. 

Figure 1. Structural features of dog calnexin, mouse calmegin, and rabbit calreticulin. Segments of sequence similarity are represented by the white boxes and the two tandemly repeated sequence motifs are indicated by the numbers 1 and 2. Sites of high affinity calcium binding (Ca +), oligosaccharide binding, and phosphorylation by casein kinase II (C.K. II) are shown. ER localization sequences are indicated at the C-terminus of each protein.

Fliegel, L., Burns, K., MacLennan, D.H., Reithmeier, R.A.F., and Michalak, M. Molecular Cloning of the High Affinity Calcium-binding Protein (Calreticulin) of Skeletal Muscle Sarcoplasmic Reticulum/. Biol Chem. 1989 264, 21522-21528. 

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