Protein Binding Affinity & Specificity

Question 1

The ____ and ____ of an interaction determines how likely the binding is to occur in the cell.

Answer 1

affinity; specificity

Question 2

The relative affinity of a paticular interaction with respect to other possible interactions.

Answer 2

specificity

Question 3

What does this picture depict?

Answer 3

left: specific binding
right: nonspecific binding

e.g. pTyr antibody binding

Question 4

Because the cellular environment is extremely concentrated with diverse proteins, any potential binding reaction is in constant ____ with alternative possibilities of interactions.

Answer 4

competition

Question 5

How does the cell allow specific interactions?

Answer 5

specificity and affinity
The higher the affinity of the binding, the more likely a specific interaction will take place.

Question 6

The ____ the affinity of the binding, the ____ likely specific interaction will take place

Answer 6

higher; more

Question 7

Depending on the context, the cell must always have ways to ____ specific context to make the interaction specific.

Answer 7

regulate

Question 8

Which protein domain binds to pTyr?

Answer 8

SH2

Question 9

The measure of the intrinsic strength of the binding interaction.

Answer 9

affinity

Question 10

The quantitative strength of a binding interaction is defined by the ____.

Answer 10

Kd - dissociation constant

Question 11

What is the simple binding reaction equation?

Answer 11

A + B <–> AB

Question 12

For stronger binding, would you predict Kd to be a higher or lower value?

Answer 12

lower

Question 13

Kd = K(off)/K(on) = ??

Answer 13

[A][B]/[AB]

Question 14

From [A][B]/[AB], how can Kd be determined?

Answer 14

Kd can be determined by the concentrations of B and AB. A eventually runs out so that is why it plateaus.

Question 15

What constant relates to the concentration of the substrate needed to make the enzyme work efficiently?

Answer 15

Km (Michaelis’ constant)

Question 16

What is defined as the concentration of a substrate when the reaction rate is half maximal, a direct measure of how tightly the substrate is bound to the enzyme?

Answer 16

Km

Question 17

How is Kd different from Km?

Answer 17

Kd is the dissociation constant, the greater it is, the less affinity it has for the substrate.
Km takes into account the rate constants for the entire reaction.

Question 18

What does a Kd of 10^-15 M tell us?

Answer 18

The interaction is strong

ex. Avidin-biotin binding

Question 19

What does a Kd of 10^-7 M tell us?

Answer 19

The interaction is semi-weak

ex. SH2 domain-phosphotyrosine (pTyr) site

Question 20

What is the thermodynamic relationship of the dissociation constant?

Answer 20

Relative amounts of free components of the complex present at equilibrium are directly related in the free energy of the two states.

Question 21

The dissociation constant is related to thermodynamics in that…
Formation of the complex will be favored if the complex has ____ energy; if the complex has higher free energy than the component, ____ amount of complex will be present.

Answer 21

lower energy; small amount

Question 22

What is the term for the idea that the world is attracted to lower energy?

Answer 22

energy minima

Question 23

What is deltaGº?

Answer 23

intrinsic free energy of the binding

Question 24

What should deltaGº be in order for protein binding to occur?

Answer 24

It must be negative (-deltaGº)
< 0

Question 25

How can mutations affect binding?

Answer 25

Small changes in binding energy will cause large difference in Kd. Thus, mutations in binding energy can change Kd.

Question 26

Which binding would be preffered for protein P?

Answer 26

The binding to B would be more favored because it generates lower energy

Question 27

The amount of time it will take for the half life of the complex to dissociate.

Answer 27

half-life

Question 28

What are the assumtions made about a theoretical calculation based on the equation?

Answer 28

1) all components are well mixed and freely diffusing in solution, which may not be always the case inside the cell
2) only one molecule of A binds to one molecule of B

Question 29

What is the effect where the presence of multiple binding sites would significantly increase the affinity?

Answer 29

Avidity effect

Question 30

Ideal affinity and specificity depends on biological function and ligand concentration.
How does the cell control this?

Answer 30

kinases and phosphatases

Question 31

What could be an advantage to lower affinity?

Answer 31

The protein can adapt to change, more interactions can occur, it is transiet and short-lived

Question 32

Which of these associations is most likely a heat shock protein?

Answer 32

hsp70/clients
-ATP -> +ATP

Question 33

Why would the hormone/receptor association have a high affinity and specificty?

Answer 33

Because the endocrine system is highly specialized and requires a direct signal to be transmitted to a direct receptor for proper processes to occur in the body. Required to be very specific and be attracted to receptor.

Ex. hypothalmus to ovaries