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BMS 242 - Core Phys/pharm > Receptor theory II > Flashcards

Receptor theory II Flashcards

1
Q

What is the binding of an agonist governed by?

A

The rate of the forwards and backwards reaction of binding

Affinity - balance between the 2

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2
Q

What is affinity and what it a balance of?

A

A measure of how well the drug binds to its receptor

The balance between the forwards and backwards reaction
(high affinity, high forwards reaction and slow reverse reaction)

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3
Q

What is occupancy a measure of and what is it dependant on

A

How many receptors are occupied by a drug at any moment in time

Dependant on AFFINITY for the drug

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4
Q

What is an antagonist?

A

A drug which binds to a receptor, without switching on receptor signalling - doesn’t directly

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5
Q

What drugs does affinity apply to?

A

Equally applies to agonists and antagonists

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6
Q

What is affinity constant for?

A

A specific drug with its specific receptor in ANY tissue

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7
Q

What does efficacy govern?

A

Activation

The change in the cell, by causing the stabilisation of the active agonist/receptor configuration

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8
Q

What does efficacy apply to?

A

Agonists only

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9
Q

On what scale is efficacy measures?

A

On a scale of 0-1

High efficacy is a value of 1

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10
Q

In the case of an agonist, how is affinity related to efficacy?

A

They are in dependant of each-other

Reactions can have a high affinity and a low efficacy and vice versa

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11
Q

What is the efficacy of an antagonist?

A

0 - Unable to induce an active configuration

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12
Q

What is occupancy a measure of?

A

The proportion of receptors occupied with a certain drug concentration

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13
Q

On what scale is occupancy measured?

A

0-1

1 = all receptors occupied
0 =no receptors occupied

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14
Q

What is occupancy NOT proportional to?

Why?

A

The size of the response

Because the maximum response of a tissue can occur with less than 100% occupancy - the tissue can have spare receptors

Maximum response of a tissue can occur at <5% occupancy

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15
Q

Why is measuring the response not a good way to measure occupancy in order to calculate affinity?

A

Size of the response is governed by both:

  • Affinity
  • Efficacy in agonists

(not just affinity)

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16
Q

What assay is used to measure occupancy to get an idea of affinity?

A

Radio ligand binding assay

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17
Q

What does a radio ligand binding assay measure?

A

Biding of a ligand (agonist or antagonist) to a protein target

Measure affinity

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18
Q

What is the main negative point of a radio ligand binding assay?

How is this overcome?

A

NONE-SPECIFIC BINDING occurs

  • Overcome by doing 2 experiments in parallel
  • Slowly increase concentration of ligand in each tube
  • Had a HUGE excess to one of the tubes
  • Displaces the specific binding and adds to the non-specific binding

Then:
Tube 1 - Radio active drug is specific and non-specific

Tube 2- Radio active drug is ONLY NON-SPECIFIC

SO tube 1 -tube 2 is specific binding of the radioactive drug

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19
Q

When choosing the radio-ligand, what must be considered?

A
  • Purity (consider isomers)
  • Degradation
  • How to label drug
20
Q

What 4 steps can be taken to minimise the degradation of the radioactive drug?

A

1) Low temperature
2) Add oxidant (minimise oxidation)
3) Avoid light (can cause chemical changes)
4) Add free-radical scavengers

21
Q

Example of a free-radical scavenger?

A

Ethanol

22
Q

Example of an antioxidant?

A

Asorbic acid

23
Q

What are 3 advantages of Tridium as a radioactive substance?

A
  • Doesn’t alter properties
  • High specific activities
  • Long half-life (good stability)
24
Q

What are 3 disadvantages of Tridium as a radioactive substance?

A
  • Dangerous to humans
  • Specialist labs needed (expensive)
  • Amount of label per molecule differs
25
Q

What are the 2 advantages of using radioactive iodine as a radioactive substance?

A
  • Easy and cheap

- Can be incorporated at high specific activities

26
Q

What are the 2 disadvantages of using radioactive iodine as a radioactive substance?

A
  • Short half-life

- Biological activity of the ligand measured can be reduced

27
Q

Why must the tissue be incubated?

A

When blend cells to release receptors - breaks up lysosomes and releases enzymes

  • Incubating prevents enzymes from breaking down the proteins and receptors
28
Q

How separate bound ligand (specific or non-specific) from non bound?

What are the problems with this method?

A

Centrifugation - Forms a pellet containing the protein

Problems:
- Specific binding may occur to the pellet (must rinse)

All these steps take time:

  • If the drug has a low affinity, off rate is quick
  • When washing pellet, may wash off some previously bound ligand
29
Q

What is Kd and how does it relate to affinity?

A

Kd - binding coefficient

The higher the Kd, the lower the affinity
Weaker the ligand is bound

30
Q

What is the shape of the non-specific binding curve?

A

Linear

31
Q

What is the shape of the specific binding curve?

What does this show?

A

Increase rapidly to a peak
Then level off as reach a maximum

Shows saturation

32
Q

How are specific binding graphs plotted and why?

A

X axis on a semi-log scale

Fit more concentrations on a reasonably sized graph

33
Q

What does the Langmuir equation describe?

What is it a plot of?

A

Describes the relationship between drug concentration and occupancy (specific binding –> therefore affinity)

On the X axis - concentration of drug (on a log scale)

On the Y axis - Specific binding

34
Q

How do you work out the Kd and Bmax from the Langmuir plot?

A

Bmax is the the level at which the graph plateaus

Kd is the measure at 50% occupancy of the total number of receptors

35
Q

What 3 things is the amount of bound ligand dependant on?

A

1) Bmax
2) Concentration of the drug
3) Kd (Binding coefficient)

36
Q

How is the Kd related to the same receptor in different tissue?

A

It is the same value if it is the same receptor-ligand match but in different tissues

As Kd is a measure of affinity

37
Q

What does Kd detetmine

A

Affinity

The on/off rate of the ligand to the receptor

38
Q

What is the scatchard equation?

A

B/F =(Bmax-B)/KD

B = specifically bound radioligand 
F = free radio ligand ( ~ concentration added)
Bmax = maximum no. of binding sites 
KD = equilibrium binding constant (affinity)

OR
B/F = - 1/Kd (B- Bmax)

39
Q

What is the scatchard plot and why is it useful?

A

Rearragement of the Langmuir equation - to make a linear plot

Better to calculate Kd

40
Q

Describe to scatchard plot

A

On the X axis:

  • Specific bound radioligand
  • X intercept is the Bmax

On the Y axis:
- B/F (bound over free radioligand)

Downwards linear plot where the slope is -1/KD

41
Q

How to calculate the Kd from the slope in the scatchard plot?

A

Slope = Y/X

Slope = -1/Kd

Kd= -1/slope

42
Q

Why is the ‘free concentration of drug’ the same as the concentration of drug added in the first place?

A

No. of drug molecules bound to receptors is insignificant

43
Q

What is the Bmax and how is this useful?

A

The maximum amount of receptors in the membrane which the drug can bind to

  • Can compare the Bmax in health and disease (is there a decrease in receptors?)
44
Q

What is the relationship between Kd and affinity?

A

Inverse relationship

Low Kd - high affinity (don’t need much of the drug)

High Kd - low affinity (need lots fo the drug to cause a response)

45
Q

What is the shape of a langmuir plot?

A

Sigmoidal

46
Q

How can you work out the Kd from a langmuir plot?

A

50% occupancy

47
Q

What can you work out from the scatchard equation?

A

Kd (and therefore the affinity of the agonist)

Bmax

BMS 242 - Core Phys/pharm (13 decks)

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