creatine kinase Flashcards
In what tissues is CK present at high levels?
Present in all cells at low levels but found in brain, heart and skeletal muscle at very high levels
When and why is CK found in the blood?
Damage to the cell membrane allows leakage of CK into the blood stream - e.g. from cell death
Why does ATP deficiency kill cells?
The pumps need ATP to maintain gradients or osmotic loss/gain. In an ischaemic cells there is a lack of ATP, change in concetration gradients and accumulation of oxygen free radicals.
How might you determine CK activity?
Use a coupled assay
Why might the three isoenzymes be separated by electrophoresis?
They have different charges so isoelectric focusing can be used. The enzymes are placed in a gel with positive charge on one side and negative on the other and the enzyme will move to where is has a net charge of 0 (isoelectric point). There is a pH gradient across the gel
(can’t use size as molecular weights are the same)
How might one establish a diagnosis of myocardial
damage?
elevated levels of MB CK in the serum
Does an increase in serum CK activity relate to the size of myocardial damage?
Yes - the levels are directly proportional to the amount of cell death in the heart as each myocyte carries an equal volume.
What is the time course of serum CK after a myocardial infarction?
30 mins to 2.5 days
What other markers can be used for diagnosis of
myocardial damage?
Lactate Dehydrogenase (LDH) - leaks out when cells are damaged. It is not particularly specific and only peaks after 6 days.
Troponin - there is a specific myocardium version - look for elevated levels of Troponin I and Troponin T which are specific to cardiac muscle (appears after 48 hours and lasts 5 days).
Serum Glutamate Oxaloacetate Transaminase (SGOT) - starts being released from cells. Peaks as CK goes down.
What are the three isoenzymes and where are they found?
BB (brain)
MM (muscles)
BM (heart)
Creatine kinase is demeric
How can increased CK be related specifically to the heart?
CK is a dimer coded for by two separate genes. They make two different isoforms - B and M. They have the same molecular weight by differ in their isoelectric point. BM form must be detected in the blood.
How else can the three isoforms be separated?
column chromatography
How can CK be tested for quickly and cheaply in hospitals?
Use immunodetection - specific antibody made to the BM isoform
How can a coupled assay be used to detect CK levels (not specific to isoform)?
creatine phosphate + ADP –> creatine and ATP
D-glucose + ATP –> ADP + G6P
G6P + NADP+ –> 6-PG + NADPH + H+
production of NADPH can be monitored by UV absorbance
What causes the plasma membrane of myocardial cells to become leaky?
Active transport membrane proteins (pumps) stop working because they require ATP to function. High concentrations of everything inside the cells leaks out.
