This is a student webpage made for an assignment at Davidson College in Dr. Campbell's Immunology Course
CHIME image of Calmodulin obtained . This file was downloaded from University of California:
Division of Biological Sciences - permission pending
Calmodulin: Biological Basics
Calmodulin is a highly conserved 17 Kd protein containing 148 amino acids, located within the cytoplasm of all cells. Calmodulin is a calcium binding protein possessing four EF-hand binding sites, which refer to calcium binding motifs found in several other proteins: calcinurin, myosin, and troponin to name a few ( Brody, 1996). Calmodulin is characterized by two domains, connected by an alpha-helix chain. Each domain has the capacity to bind two calcium ions. As stated above calmodulin possess two EF-hands, or binding domains, giving it the ability to bind a total of four calcium ions. Calmodulin in its unbound state has a dumbbell conformation. Binding Ca2+ ions causes a conformational change in calmodulin, making it available for interaction with target proteins( Brody, 1996) . The gene that produces calmodulin produces two transcripts of 1.6 and 1.7 Kb in addition to containing 5 exons( Brody, 1996). Calmodulin is a commonly known intracellular calcium receptor that functions in the regulation of biological activities of various proteins within cells. It also plays a role in transmembrane ion transport ( Brody, 1996) . Calcium ions serve as an activator of calmodulin, inducing conformational changes, which alls the changed protein to bind target proteins. By binding theses specific proteins calmodulin calmodulin is able to initiate various signaling cascades that are important in immune system function. Calmodulin is involved in most of the important signaling pathways in cells. Some of its targets include: cyclic nucleotide metabolism, phosphorylation pathways, dephosphorylation, and calcium transport ( Brody, 1996).
Fig 2: Homo Sapien amino acid sequence click here permsission pending-
Fig 3: Side / center views of Calmodulin bound to Ca2+ ions. click here permission pending
Anthrax and Calmodulin:
Anthrax produces three toxins that are necessary for destroying targeted cells: protective antigen, lethal factor, and edema factor. Protective antigen is responsible for clearing a path through cellular membranes so that the other two toxins can enter the cell. Lethal factor is responsible for destroying cells belonging to the immune system. The third toxin, edema factor, is of particular interest because of its interaction with the protein calmodulin. Edema factor is responsible for fluid accumulation, and causes death by causing excess fluids to be released into lungs or other infected areas. This fluid accumulation serves to make lethal factor, the immune cell destroying toxin of Anthrax, ~ 100x stronger. Edema factor mimics the functions of cyclic AMP (cAMP), causing cells use reserved energy more quickly, and causing them to lose the ability to regulate their environments. This in turn causes water to be released and the cells to die. The curious phenomenon surrounding edema factor is that it is totally benign upon entering the cell. Only when interacting with the protein calmodulin does edema factor become active. Binding calmodulin activates edema factor by causing a conformational change in the toxin that causes it to closely resemble and behave like cAMP.
Fig. 4 shows Edema Factor (EF) by itself and after activation with Calmodulin (CaM). permission pending
Parkinson's Disease and Calmodulin
Parkinson's disease is a neurological disease affecting nerve cells responsible for controlling muscle movement. Symptoms include trembling, muscle rigidity, problems with coordination and balance (Mayo Clinic, 2003). The disease is progressive, and become disabling over time. Loss of cells in an area of the brain called the substantia nigra (part of the brain stem) is associated with the onset and progression of Parkinson's Disease (Lieberman, 1996). Loss of cells in the substantia nigra is restricted to those cells that contain dopamine. Although many cells in the brain contain dopamine, those affected by Parkinson's Disease lack the Ca2+ ion dependent protein calmodulin (Lieberman, 1996). Those cells expressing both dopamin and calmodulin survive, while those cells lacking calmodulin die prematurely, leading to the onset of Parkinson's disease (Lieberman, 1996).
Fig. 3: Shows absence of substantia nigra in brains
with Parkinsons in comparison with normal brains: th
R = red nucleus, sn = substantia nigra, lgn = lateral genicalute body. permission pending
Rheumathid Arthritis and Calmodulin
T-cell dependent immune mechanisms are thought to be closely associated with the onset o rheumatoid arthritis. Rheumatoid Synovial T-cells associated with this disease bear some resemblance to anergic T-cells in that they exhibit characteristics such as hyporesponsiveness and poor proliferation (John Hopkins, 1998). An experiment comparing transcription of anergy related genes in tissues from patients with RA and those with Reactive Arthritis, revealed that three genes were downregulated in those patients with RA compared to reactive arthritis (John Hopkins, 1998). The three downregulated genes were cellular apoptosis susceptibility protein, BF626661, and Calmodulin. Transcription of calmodulin in patients with Rheumatoid arthritis was less than 1% the amount of calmodulin transcribed in Reactive Arthritis (John Hopkins, 1998). Researchers suspect that reduced transcription of the protein calmodulin may play a role in the hyporesponsiveness and apoptotic defect of RA T-cells. Patients were treated with a drug called Infliximab, which upregulated production of calmodulin by approximately 10x its previous expression (John Hopkins, 1998). Although it does not bind calmodulin directly, infliximab plays an important role in upregulating production of calmodulin in patients with rheumatoid arthritis. Infliximab has been found to be affective in certain stages of Crohn's disease as well as in rheumatoid arthritis (Atena, 2003). It is a monoclonal antibody that binds and TNF-alpha, which is responsible for cell death in the face of chemical or physical injury. Inhibition of TNF-alpha allows for the upregulation of calmodulin (Atena, 2003).
R e f r e n c e s
Atena (2003) Coverage Policy Bulletins: Remicaid (Infliximab) 14 Feb. 2003 http://www.aetna.com/cpb/data/CPBA0341.html
Brody, T.B., (1996) Calmodulin: Biological Overview. 1 Feb 1997. http://flybase.bio.indiana.edu/allied-data/lk/interactive-fly/dbzhnsky/calmod1.htm
John Hopkins (1998) Transcription of Several Genes Associated with Antigen Induced Anergy are Regulated by Rheumatoid Arthritis Synovial T-cells. 2001. http://www.hopkins-arthritis.som.jhmi.edu/news-archive/2001/anergy.html
Kim, J., (2002) Calmodulin Target Database: Binding Site Search and Analysis. NA. http://calcium.uhnres.utoronto.ca/ctdb/ctdb/home.html
Mayo Clinic (2003) Brain and Nervous System Center. 18 July 2002. http://www.mayoclinic.com/invoke.cfm?id=DS00295
Lieberman, A.(1996). Understanding Proteins, Understanding Parkinson's Disease. 2003. http://www.parkinson.org/protein.htm
Protein Music. (2002) Calmodulin. 15 Aug. 2001. http://www.whozoo.org/mac/Music/Calmod.htm
University of California: Division of Biological Sciences.(2002) Images of Biological Molecules. NA. http://www-biology.ucsd.edu/classes/bibc100.FA98/images.html
University of Chicago Hospitals (2003) Structure of Anthrax Toxin Offers Clues to Treatment. 23 Jan 2002. http://www.uchospitals.edu/news/2002/20020123-efactor.html