Creatine Kinase 5 Isoforms Know in the Body Cystosolic (70% of enzymatic activity) 1.) Muscle type (CK-MM) -homodimer 2.) Brain type (CK-BB) 3.) Hybrid type (CK-MB) -heterodimer -typically found in the heart Mitochondrial (30% of enzymatic activity) 4.) Sarcomeric type (smtCK) -octamer 5.) Ubiquitous type (umtCK) Blum, H. E., Deus, B., and Gerok, W. (1983) Mitochondrial creatine kinase from human heart: purification and characterization of crystallized isoenzyme. J. Biol. Chem. 94, 1247-1257. Creatine kinase is an enzyme that catalyzes the phosphorlatyion reaction provided by the image. It provides an energy buffer in times of stress. The equilibrium is constantly at work though. Phosphocreatine also provides an energy shuttle that can transport energy between subcellular units. CK-BB is found in non-skeletal muscle tissue but is mainly localized in the brain. CK-MB has monomeric units from both CK-BB & CK-MM units. smtCK is only found in mitochondria of skeletal muscle. Ubiquitous found in all other. 30% of all creatine kinase activity is to be attributed to mitochondrial.

Pathway Tokasarska-Schlattner, M., Williann, T. (2006) Mitochondrial creatine kinase in human health and disease. Mol Bas. Dise. Vol 176, Iss 2, 164-180.

Amino Acid Sequence Alignment -Methionine 273 -Cysteine 284 Black shaded amino acids are considered completely conserved amino acids. Gray shaded sections represent aligned amino acids that are not identical, but have similar characteristics. Creatine kinase has strong conservation over all isoforms making it a very stable protein. The highlighted residues are functionally important. These numbers assigned to the the specific amino acids may vary between isoforms, but they will still align properly. Met267 is important for formation of octamer and dimer. Cys284 is very important to the binding of the substrate. Its deprotanted negative charge also helps in active site structure. Try these residues provide a pocket for the binding site. Lyzlova, S. N., Stefanov, V. E. (1990) Phosphagen Kinases. CRC Press, 63-73 pgs. -Tryptophan 211, 218, and 228

Structure -Human CK-BB -Contains -19 alpha helices (38%) -18 beta sheets (16%) -13-18 alpha helicies depending on specific structure -32-38% alpha helix residues -16-19 beta sheets -15-16% PDB id = 3DRB Bong, S. M., Chi, Y. M., Hwang, K. Y., Lee, K. S., Moon, J. H., and Nam, K. H. (2008) Structural studies of human brain-type creatine kinase complexed with ADP-Mg2+-NO3—creatine trasition-state analogue complex. FEBS Lett. 582, 28, 3959-3965. This unit is from the cytosol of a human brain cell. This particular structure contains 19 helices and 18 beta sheets. This is not consistent across the board. From the solved structures in the Protein Data Bank the structures range from 13-18 helices and 16-18 beta sheets.

Important Residue Interaction -Red loop 59-69 -Purple His65 -Green loop 320-330 -Purple Asp325 -Yellow Cys283 Forest, E., Marcillat, O., Mazon, H. and Vial, C. (2003) Changes in MM-CK conformational mobility upon formationof the ADP-Mg2+-NO3—Creatine transition state analogue complex as detected by hydrogen/deuterium exchange. J. Biochem. 42, 13596-13604. This model shows the above residues…….The interaction of these looped regions is vital to the success of this protein. The yellow cysteine represents the approximate location of the active site. As the substrate binds to this region the two colored loop regions will close around it. The highlighted purple residues bind with each other to form a water excluding bond. What is interesting is that this conformational change will not take place through the binding of only creatine or phosphocreatine. There was a study done (specifically on CK-BB) that isolated different stages of the binding process.

Transition Complex -Green loop interactions close off active site -Arg 130,132,236,292,320 create a positive pocket for phosphate of ATP/ADP to bind -Creatine’s carboxylate forms a H-bond with main chain N from Val72 -Cys283 anchors creatine for nucleophilic attach of a phosphorus from ATP Bong, S. M., Chi, Y. M., Hwang, K. Y., Lee, K. S., Moon, J. H., and Nam, K. H. (2008) Structural studies of human brain-type creatine kinase complexed with ADP-Mg2+-NO3—creatine trasition-state analogue complex. FEBS Lett. 582, 28, 3959-3965.

Creatine Kinase Test -Typical cardiac marker - During myocardia infarction cell contents is expelled -Can be an indicator of several other health issues -Depends on what isoform is elevated in the blood -Stroke, pulmonary infection, muscular dystrophies, myopathy, and others "Creatine Kinase, Serum." Mayo Clinic Mayo Medical Laboratories. January 1, 1999. Accessed October 19, 2014. http://www.mayomedicallaboratories.com/test-catalog/Clinical and Interpretive/8336.

Creatine Supplementation -Can be utilized for short burst of energy for athletes -Strength and Endurance -Parkinson’s (slow symptoms) -Precautions??? "Creatine." Medline Plus. September 8, 2014. Accessed October 19, 2014. http://www.nlm.nih.gov/medlineplus/druginfo/natural/873.html.