Creatine Kinase

Question 1

What is creatine kinase used for?

Answer 1

To convert creatine phosphate into Creatine and ATP

Question 2

What is delta G hydrolysis of ATP and creatine phosphate?

Answer 2

-31kJ/mole for ATP and -43.1kJ/mole for creatine phosphate

Question 3

What is a blood test for usually, creatine phosphate, creatine or creatine kinase?

Answer 3

creatine kinase

Question 4

Where in the body is creatine kinase found in particularly high concentrations?

Answer 4

in the brain and muscle cells

Question 5

When is creatine kinase released into the circulation?

Answer 5

Following death / damage of brain and muscle cells

Question 6

How many isoforms are there of creatine kinase?

Answer 6

Three isoforms/ three dimeric isoenzymes - can be identified through gel electrophoresis

Question 7

How do you conduct the gel electrophoresis?

Answer 7

on cellulose acetate strips

Question 8

What are the three isoforms of creatine kinase?

Answer 8

MM, MB and BB

Question 9

What is the only place in which the MB isoform is found?

Answer 9

Cardiac muscle/ myocardium

Question 10

How much of total creatine kinase does MB isoform represent in the myocardium?

Answer 10

about 15%, the rest is MM

Question 11

Which isoform moves closest to the negative electrode?

Answer 11

MM isoform

Question 12

What is myocardial infarct?

Answer 12

death of heart muscle cells

Question 13

why do the heart muscle cells die?

Answer 13

lack of oxygen

Question 14

Why would there be a lack of oxygen in the heart muscles?

Answer 14

Blockage of the cardiac arteries. This process is termed atherosclerosis.

Question 15

Why do cells need oxygen, how do they use it and why do cells die without it?

Answer 15

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.

Question 16

When and why is creatine kinase found in the blood?

Answer 16

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

Question 17

IN what tissues is CK present at high levels?

Answer 17

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.

Question 18

Describe the absorption spectrum for NADH and NAD+?

Answer 18

They are distinct

Question 19

What technique is used to detect CK activity in the serum?

Answer 19

coupled assay

Question 20

What is the couples assay for creatine kinase (3 equations)?

Answer 20

creatine phosphate + ADP–> creatine + ATP (via CK)

D-glucose + ATP –> ADP + G6P (via hexokinase)

G6P + NADP+ –> 6-PG + NADPH + H+ (via G6P dehydrogenase)

Question 21

How then can increased CK be related specifically to the death of cardiac muscle rather than skeletal muscle and brain tissue?

Answer 21

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.

Question 22

Which form of creatine kinase is only expressed in the brain?

Answer 22

BB isoform as the brain only expresses the B gene

Question 23

Which isoform of creatine kinase is only found in skeletal muscle?

Answer 23

MM form is the only one made in skeletal muscle cells

Question 24

What is CK?

Answer 24

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.

Question 25

Why is the level of CK BM isoform in the serum directly proportional to the amount of cell death in the heart?

Answer 25

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

Question 26

In what instance would measuring MM levels be useful?

Answer 26

to determine the extent of skeletal muscle damage in conditions like muscular dystrophy

Question 27

What is the level of MB isoform in the serum directly proportional to?

Answer 27

to the amount of cell death in the heart

Question 28

is a simple measure of CK activity in serum a sufficiently good diagnostic test for myocardial infarct”?

Answer 28

No, because the activity could be from any of three tissues. Assay of the BM form is essential.

Question 29

In general how might you experimentally discern between different protein isoforms, what methods can be used?

Answer 29

Electrophoresis or column chromatography - but are slow and require expert technical operation. Neither are ideal in a Casualty Department.

Question 30

How many days after an heart attack is CK concentration highest?

Answer 30

24-48 hours after

Question 31

What are the four markers for myocardial damage?

Answer 31

Creatine Kinase

SGOT - Serum glutamate oxaloacetate transaminase

LDH - lactate dehydrogenase

cardiac troponin

Question 32

Why is cardiac troponin useful are a marker for myocardial damage?

Answer 32

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).

Question 33

What causes the plasma membrane of myocardial cells to become leaky?

Answer 33

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

Question 34

Why might the three isoenzymes be separated by electrophoresis?

Answer 34

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)

Question 35

What is the equation for the conversion of creatine phosphate? And what is delta G of the reaction?

Answer 35