Creatine Kinase Flashcards
What is creatine kinase used for?
To convert creatine phosphate into Creatine and ATP
What is delta G hydrolysis of ATP and creatine phosphate?
-31kJ/mole for ATP and -43.1kJ/mole for creatine phosphate
What is a blood test for usually, creatine phosphate, creatine or creatine kinase?
creatine kinase
Where in the body is creatine kinase found in particularly high concentrations?
in the brain and muscle cells
When is creatine kinase released into the circulation?
Following death / damage of brain and muscle cells
How many isoforms are there of creatine kinase?
Three isoforms/ three dimeric isoenzymes - can be identified through gel electrophoresis
How do you conduct the gel electrophoresis?
on cellulose acetate strips
What are the three isoforms of creatine kinase?
MM, MB and BB
What is the only place in which the MB isoform is found?
Cardiac muscle/ myocardium
How much of total creatine kinase does MB isoform represent in the myocardium?
about 15%, the rest is MM
Which isoform moves closest to the negative electrode?
MM isoform
What is myocardial infarct?
death of heart muscle cells
why do the heart muscle cells die?
lack of oxygen
Why would there be a lack of oxygen in the heart muscles?
Blockage of the cardiac arteries. This process is termed atherosclerosis.
Why do cells need oxygen, how do they use it and why do cells die without it?
ATP is generated Via glycolysis, the Krebs Cycle and eventually oxidative phosphorylation.
The end point of the process requires atmospheric oxygen, hence if there is less oxygen supplied to a cell there is less ATP, pumps do not function, ion balance is lost and cells die.
When and why is creatine kinase found in the blood?
Cell contents are released when they are dying, e.g., proteins that should be held inside against concentration gradients appear in the serum.
Therefore the levels of many proteins including creatine kinase (many others as well such as lactate dehydrogenase) in serum can be used as indirect indicators of cell death
IN what tissues is CK present at high levels?
Creatine kinase is present in all cells at very low levels but is at high concentrations in metabolically very active tissues including the brain, heart and skeletal muscles.
Describe the absorption spectrum for NADH and NAD+?
They are distinct
What technique is used to detect CK activity in the serum?
coupled assay
What is the couples assay for creatine kinase (3 equations)?
creatine phosphate + ADP–> creatine + ATP (via CK)
D-glucose + ATP –> ADP + G6P (via hexokinase)
G6P + NADP+ –> 6-PG + NADPH + H+ (via G6P dehydrogenase)
How then can increased CK be related specifically to the death of cardiac muscle rather than skeletal muscle and brain tissue?
The only tissue where both genes are expressed is cardiac muscle cells. They therefore make all three dimers including the hybrid MB form. (only traces of BB in myocardium)
Thus, death of cardiac muscle fibres can be determined if the MB isoform of CK can be detected in the serum.
Which form of creatine kinase is only expressed in the brain?
BB isoform as the brain only expresses the B gene
Which isoform of creatine kinase is only found in skeletal muscle?
MM form is the only one made in skeletal muscle cells
What is CK?
CK is a protein made from two subunits or monomers i.e., it is a dimer.
The two monomers are coded for by two different genes,
Forming two different monomer isoforms B and M
The two monomers have approximately the same molecular mass but they differ in their pI (isoelectric point)
This means they can separate by charge.
Why is the level of CK BM isoform in the serum directly proportional to the amount of cell death in the heart?
This is because each myocyte can be considered to be approximately of equal volume (they all have equal likelihood of dying independent of their size)
So as each cell dies it releases a “quantum” of CK into the extracellular fluid and thence into the serum
In what instance would measuring MM levels be useful?
to determine the extent of skeletal muscle damage in conditions like muscular dystrophy
What is the level of MB isoform in the serum directly proportional to?
to the amount of cell death in the heart
is a simple measure of CK activity in serum a sufficiently good diagnostic test for myocardial infarct”?
No, because the activity could be from any of three tissues. Assay of the BM form is essential.
In general how might you experimentally discern between different protein isoforms, what methods can be used?
Electrophoresis or column chromatography - but are slow and require expert technical operation. Neither are ideal in a Casualty Department.
How many days after an heart attack is CK concentration highest?
24-48 hours after
What are the four markers for myocardial damage?
Creatine Kinase
SGOT - Serum glutamate oxaloacetate transaminase
LDH - lactate dehydrogenase
cardiac troponin
Why is cardiac troponin useful are a marker for myocardial damage?
Cardiac troponin I and troponin T are tissue specific and are not present in any other muscle. Their presence in the serum represents a specific marker for cardiac infarction (typically appearing in the serum after 48h of infarction and persisting for approximately 5 days).
What causes the plasma membrane of myocardial cells to become leaky?
A semi-permeable membrane separates the inside from the outside of a cell, actively excluding some things e.g. Na+ions.
This needs a protein pump in the membrane (ATPases)
They use energy in the form of adenosine triphosphate (ATP) to pump ions.
When no oxygen, no oxidative phosporylation, leading to no ATP, pumps stop working, homeostatic balance is lost, so things leak out the cells
Why might the three isoenzymes be separated by electrophoresis?
CK is a protein made from 2 monomers = it is a dimer
The two monomers have approximately the same molecular mass but differ in their pI (isoelectric point).
This means that they can be separated by charge)
What is the equation for the conversion of creatine phosphate? And what is delta G of the reaction?
