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Calmodulin (CaM) is approximately 148 amino acids long (Chattopadhyaya et al., 1992). For more information about the amino acid primary sequence and the protein's secondary struture use the embedded links). It is a dumbbell shaped protein with two calcium-binding globular domains representing the weights at either end connected by a linker sequence that is a 28 amino acid long alpha helix (Finn et al., 1995). The below picture is a diagram of the calmodulin protein with 4 calcium ions (pink balls) bound to the globular domains and a light blue alpha helix connecting the two regions.
(Dutta et al., 2003)
The calmodulin protein contains 4 EF hand motifs, 2 at each globular calcium binding domains. The EF-hand motif is highly conserved amongst various calcium binding proteins. Each EF hand motif supplies an electronegative environment suitable for 1 calcium ion to bind (Dutta et al., 2003). The below illustration demonstrates what an EF-motif looks like and how it creates a "pocket" for a calcium ion to bind:
(Biochemistry 462a: Enzyme Regulation, 2003)
The two symmetrical globular domains also known as lobes contain three alpha helcies and two calcium ion binding EF hand motifs, which are separated by a short beta-shee (Chattopadhyaya et al., 1992). The two lobes are separated by a flexible alpha helix linker sequence, which allows the two globular domains to move and bend. The flexibility of this linker alpha helix will be explained later in the following section.
There are four main aspects of calmodulin's structure that allow for the versatile functionality of the protein within calcium induced signal transduction pathways:
1). The EF-hand motifs
The existence and separated EF-hand motifs on the calmodulin protein allow for calcium ions to bind and therefore allow for the protein to sense an increased presence of calcium. Without these binding domains, calcium ions could not bind to the protein and the protein could not carry the calcium induced signal transduction pathway.
2). The location of non-polar regions and the conformational change when calcium ions bind
Once calcium ions have bound to the calmodulin protein structure at all 4 EF-motifs, the protein undergoes a conformational change. After calcium binds hydrophobic (non-polar) methyl groups from methionine amino acid residues become exposed on the protein. These non-polar R groups from methionine where hidden within the structure before the conformational change, and it was the binding of the calcium ions that exposed these regions. It is important to note that in calmodulin, methionine is an unusually abundant amino acid (Dutta et al., 2003).
The below picture indicates the conformational change that occurs when 4 calcium ions (light blue circles) bind to calmodulin. The nonpolar regions are colored with green carbon atoms and the methionin sulfur atoms are in yellow, indicating that these regions are now exposed, whereas before the calcium binding (the picture on the left) these regions where hidden. Note that the non-polar amino acids form "grooves" (indicated with red stars), where calmodulin can bind to and activate other proteins. Also notice that these "grooves" are generic in shape, making calmodulin a protein that is able to bind to a variety of target proteins with a variety of shapes and structures. Also the fact that the interaction between calmodulin and its target protein rely on the interaction between non-polar domains allows for more versatility in which target proteins calmodulin can bind to. For instance, calmodulin does not require a particular amino acid sequence on the target protein in order to bind, but only requires a motif of non-polar basic amino acids (Dutta et al., 2003). The fact that calmodulin does not require a specific amino acid sequence to bind to a target protein may leave some skeptical about the strength of the protein-protein interaction, but the flexible linker sequence helps strengthen the protein -protein interactions between calmodulin and other target proteins.
(Dutta et al., 2003)
3). The felxibility of the linking alpha helix sequence
The two symmetrical domains with the calcium-binding EF-hand motifs are separated with a flexible alpha helix linker sequence (Dutta et al., 2003). The flexibility of this sequence allows calmodulin to wrap around its target protein, which helps form a more stable connection between calmodulin and the protein is is activating. The illustration below shows two examples of calmodulin binding to two different target proteins shown in red (top: calmodulin-dependent protein kinase II-alpha and bottom: myosin light chain kinase) and how the flexible linking sequence (purple) can wrap around different target molecules. The diagram also shows that the flexible linking sequence (purple) allows for versatility in terms of what target molecules calmodulin can bind to because it can readjust based on the structure of the specific target protein (Dutta et al., 2003). In other words the diagram shows that depending on the target protein, the linking sequence can move and change conformation, therefore being felxible depending on the target protein. Hence, the flexibility of the linking sequence allows for calmodulin to form a stronger protein-protein interaction and allows for calmodulin to bind to various different target protein structures.
(Dutta et al., 2003)
4). The placement of amino acids able to be phosphorylated
Finally, researchers also believe that calcium binding is not the only mechanism by which the calmodulin can be activated and controlled. Researchers also think that the mechanism of phosphorylation may help modulate calmodulin activity (Sacks et al., 1995). The placement of amino acids that are able to be phosphorylated (such as serine and threonine) in sites that are accessible to kinases is another structural component of calmodulin that allow for the regulation of its function.
In conclusion, the structure of calmodulin, including the placement and existence of EF-hand motifs, the flexibility of the linking alpha helix sequence, the conformational changes and placement of non-polar interaction residues after calcium binding, and the existence of amino acids able to be phosphorylated allow for the calmodulin protein to act as a calcium-binding intermediate in signal transduction pathways and accounts for the ability of calmodulin to bind to and activate a variety of target proteins.
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