Calcium binding/EF hands

The calcium binding proteins calbindin D28k, calretinin and parvalbumin are members of a family of proteins characterized by the presence of calcium binding EF-hand motifs, modulated by stimulus-induced increases in cytosolic free calcium ions (Persechini et ah,... 

When proteins fold into their tertiary structures, there are often subdivisions within the protein, designated as domains, which are characterised by similar features or motifs. A protein domain is a part of the protein sequence and structure that can evolve, function and exist independently of the rest of the protein chain. Many proteins consist of several structural domains. One domain may appear in a variety of evolutionarily related proteins. Domains vary in length from about 25 up to 500 amino acids. The shortest domains, such as zinc fingers , are stabilised by metal ions or disulfide bridges. Domains often form functional units, such as the calcium-binding EF hand domain of calmodulin. As they are self-stable, domains can be swapped by genetic engineering between one protein and another, to make chimera proteins. 

The structures also revealed a Zn2+-binding site in the protein. The coordination of the zinc ion is similar to the pattern seen in certain metalloproteases (Brodersen et al 1999). Sequence comparison suggests that this zinc-binding site is present in a number of the remaining members of the S100 family (Fritz and Heizmann, 2004). The structure of S100A7, crystallised in the absence of zinc, further showed that loss of zinc resulted in a reorganisation of the adjacent empty and distorted EF-hand loop, similarly to a calcium-loaded EF-hand. 

Parvalbumin is a muscle protein with a single polypeptide chain of 109 amino acids. Its function is uncertain, but calcium binding to this protein probably plays a role in muscle relaxation. The helix-loop-helix motif appears three times in this structure, in two of the cases there is a calcium-binding site. Figure 2.13 shows this motif which is called an EF hand because the fifth and sixth helices from the amino terminus in the structure of parvalbumin, which were labeled E and F, are the parts of the structure that were originally used to illustrate calcium binding by this motif. Despite this trivial origin, the name has remained in the literature. 

Figure 6.21 Schematic diagram of the conformational changes of calmodulin upon peptide binding, (a) In the free form the calmodulin molecule is dumhhell-shaped comprising two domains (red and green), each having two EF hands with bound calcium (yellow), (b) In the form with bound peptides (blue) the a helix linker has been broken, the two ends of the molecule are close together and they form a compact globular complex. The internal structure of each domain is essentially unchanged. The hound peptide binds as an a helix.

Figure 6.21a) comprising two domains separated by a long straight a helix, similar in shape to troponin-C described in Chapter 2 (see Figure 2.13c). Each domain comprises two EF hands (see Figure 2.13a), each of which binds a calcium atom. The two domains are clearly separated in space at the two ends of the a helix linker. 

These cytosolic proteins contain five EF-hand domains and are able to translocate to the plasma membrane upon calcium binding [5]. In addition to the EF-hand domains, these proteins also have a hydrophobic glycine/proline-rich domain, important for their translocation to the membrane. To date five members of this... 

ALG-2 is the fust calcium-binding protein of the EF-hand family found to be directly involved in apoptosis. ALG-2 is a 22 kDa protein and like the other members of the penta EF-hand family, contains five EF-hands, with only two of them functional. ALG-2 protein is expressed in the brain and eye and was found to be upregulated in various cancer tissues. Several targets have been found, such as proteins AEP, Alix, preflin, and annexins, suggesting a putative role of ALG-2 in apoptosis. 

Krebs J, Heizmann CW (2007) Calcium-binding proteins and the EF-hand principle. In Krebs J, Michalak M (eds) Calcium a matter of life and death, New Comprehensive Biochemistry, vol 41 (Series Bernardi G). Elsevier, pp 51-93... 

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).

The EF-hand construct of calcium-binding proteins in cellulases (prokaryotes), but only single hands, becomes extended to triggers (eukaryotes) in twin hands. 

Many calcium-binding proteins can be described in terms of loops containing potential metal-binding sites linked by helices of various shapes and lengths. There are several families of closely related proteins, in particular the groups of EF-hand species and... 

Obelin is a Ca2+-activated bioluminescent photoprotein that has been isolated from the marine polyp Obelia longissima. Binding of calcium ions determines a luminescent emission. The protein consists of 195 amino acid residues [264] and is composed of apoobelin, coelenterazine, and oxygen. As aequorin, it contains three EF-hand Ca2+-binding sites and the luminescent reaction may be the result of coelenterazine oxidation by way of an intramolecular reaction that produces coelenteramide, C02, and blue light. As for aequorin, the luminescent reaction of obelin is sensitive to calcium and the protein was used in the past as an intracellular Ca2+ indicator. More recently, the cloning of cDNA for apoobelin led to the use of recombinant obelin as a label in different analytical systems. 

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