Many calcium-binding proteins belong to the same evolutionary family and share a type of calcium-binding domain known as the EF-hand [1,2,3,4,5]. This type of domain consists of a twelve residue loop flanked on both side by a twelve residue α-helical domain (see <PDB:1CLL>). In an EF-hand loop the calcium ion is coordinated in a pentagonal bipyramidal configuration. The six residues involved in the binding are in positions 1, 3, 5, 7, 9 and 12; these residues are denoted by X, Y, Z, -Y, -X and -Z. The invariant Glu or Asp at position 12 provides two oxygens for liganding Ca (bidentate ligand). The basic structural/functional unit of EF-hand proteins is usually a pair of EF-hand motifs that together form a stable four-helix bundle domain. The pairing of EF-hand enables cooperativity in the binding of Ca2+ ions.

We list below the proteins which are known to contain EF-hand regions. For each type of protein we have indicated between parenthesis the total number of EF-hand regions known or supposed to exist. This number does not include regions which clearly have lost their calcium-binding properties, or the atypical low-affinity site (which spans thirteen residues) found in the S-100/ ICaBP family of proteins [6].

• Aequorin and Renilla luciferin binding protein (LBP) (Ca=3).
• α actinin (Ca=2).
• Calbindin (Ca=4).
• Calcineurin B subunit (protein phosphatase 2B regulatory subunit) (Ca=4).
• Calcium-binding protein from Streptomyces erythraeus (Ca=3?).
• Calcium-binding protein from Schistosoma mansoni (Ca=2?).
• Calcium-binding proteins TCBP-23 and TCBP-25 from Tetrahymena thermophila (Ca=4?).
• Calcium-dependent protein kinases (CDPK) from plants (Ca=4).
• Calcium vector protein from amphoxius (Ca=2).
• Calcyphosin (thyroid protein p24) (Ca=4?).
• Calmodulin (Ca=4, except in yeast where Ca=3).
• Calpain small and large chains (Ca=2).
• Calretinin (Ca=6).
• Calcyclin (prolactin receptor associated protein) (Ca=2).
• Caltractin (centrin) (Ca=2 or 4).
• Cell Division Control protein 31 (gene CDC31) from yeast (Ca=2?).
• Diacylglycerol kinase (EC 2.7.1.107) (DGK) (Ca=2).
• FAD-dependent glycerol-3-phosphate dehydrogenase (EC 1.1.99.5) from mammals (Ca=1).
• Fimbrin (plastin) (Ca=2).
• Flagellar calcium-binding protein (1f8) from Trypanosoma cruzi (Ca=1 or 2).
• Guanylate cyclase activating protein (GCAP) (Ca=3).
• Inositol phospholipid-specific phospholipase C isozymes γ-1 and delta-1 (Ca=2) [10].
• Intestinal calcium-binding protein (ICaBPs) (Ca=2).
• MIF related proteins 8 (MRP-8 or CFAG) and 14 (MRP-14) (Ca=2).
• Myosin regulatory light chains (Ca=1).
• Oncomodulin (Ca=2).
• Osteonectin (basement membrane protein BM-40) (SPARC) and proteins that contains an 'osteonectin' domain (QR1, matrix glycoprotein SC1) (see the entry <PDOC00535>) (Ca=1).
• Parvalbumins α and β (Ca=2).
• Placental calcium-binding protein (18a2) (nerve growth factor induced protein 42a) (p9k) (Ca=2).
• Recoverins (visinin, hippocalcin, neurocalcin, S-modulin) (Ca=2 to 3).
• Reticulocalbin (Ca=4).
• S-100 protein, α and β chains (Ca=2).
• Sarcoplasmic calcium-binding protein (SCPs) (Ca=2 to 3).
• Sea urchin proteins Spec 1 (Ca=4), Spec 2 (Ca=4?), Lps-1 (Ca=8).
• Serine/threonine specific protein phosphatase rdgc (EC 3.1.3.16) from Drosophila (Ca=2).
• Sorcin V19 from hamster (Ca=2).
• Spectrin α chain (Ca=2).
• Squidulin (optic lobe calcium-binding protein) from squid (Ca=4).
• Troponins C; from skeletal muscle (Ca=4), from cardiac muscle (Ca=3), from arthropods and molluscs (Ca=2).

There has been a number of attempts [7,8] to develop patterns that pick-up EF-hand regions, but these studies were made a few years ago when not so many different families of calcium-binding proteins were known. We therefore developed a new pattern which takes into account all published sequences. This pattern includes the complete EF-hand loop as well as the first residue which follows the loop and which seem to always be hydrophobic. We also developed a profile that covers the loop and the two α helices.