Annexin, calcium binding sites

The first membrane binding is mediated via multiple calcium binding sites located at the convex side of the disk-shaped annexin core. The N-terminus of the protein that is presumed to harbor the second membrane binding site is buried in the core domain in the absence of Cd , and is supposed to get exposed to the aqueous phase if annexin Al binds to a membrane in the presence of calcium ions (see Fig. 6) [42]. To verify this hypothesis, again SSMs are well suited, since they allow separation of the first and second membrane binding events. N-terminal truncated annexin Al an-... 

Fig. 13. The binding sites of calcium in (a) parvalbumin (41a), (b) annexin (41) and (c) calmodulin (42). The drawings show two bidentate carboxylates coordinated to Ca2 in the EF-hand site of parvalbumin, and one bidentate carboxylate coordinated to Ca2 in annexin and calmodulin. All the donor atoms coordinated to the calciums are oxygen donor atoms from carboxylates of asp = aspartate, or glu = glutamate, or else peptide carbonyl oxygens from gly = glycine or met = methionine. Redrawn after Refs. (41-42).

Annexins isolated from chondroblast matrix vesicles may be reconstituted with phospholipids to form calcium ion channels in the complete absence of Ca2+ ions. Indeed, annexin V has domains that directly bind calcium ions glutamate and aspartate residues provide the ion binding site (EF-hand domains). Figure 9.6b illustrates putative annexin V channels that mediate an influx of Ca2+ ions into artificial bilayers and liposomes (detectable with a calcium-sensitive fluorescent dye). These in vitro annexin Ca2+ channels, and also the Ca2+ influx into matrix vesicles in cell culture and in vivo, are blocked by Zn2+ ions, or a derivative of 1,4-benzothiazepine (inhibitor K201). 

Fig. 8. Stereo view of the phospholipid binding site in the third domain of rat annexin V, including calcium ions in sites AB and AB, (a) with bound glycerophospho-serine (GPS), and (b) with bound glycerophosphoethanolamine (GPE). GPS and GPE as well as the calcium ions are drawn dark. The oxygens in the protein fragment are drawn gray. (Redrawn from Swaiijo et al. 1995).

Annexin V is a protein tliat binds to membranes in a calcium-dependent manner (Sopkova et al. 1994). In human annexin V, crystallography studies have shown that the unique Trp, Trp 187 residue, has its side chain buried in the core of the protein. It has been observed also that, in the triclinic crystal form of the protein, formation of the calcium site in the protein domain where Trp 187 is present, is accompanied by a conformational change, bringing the tryptophan side chain onto the surface of the molecule (Sopkova et al. 1993). Binding of Calcium to annexin V has then investigated by absorption and fluorescence studies. 

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