Protein Binding Affinity & Specificity Flashcards
The ____ and ____ of an interaction determines how likely the binding is to occur in the cell.
affinity; specificity
The relative affinity of a paticular interaction with respect to other possible interactions.
specificity
What does this picture depict?
left: specific binding
right: nonspecific binding
e.g. pTyr antibody binding
Because the cellular environment is extremely concentrated with diverse proteins, any potential binding reaction is in constant ____ with alternative possibilities of interactions.
competition
How does the cell allow specific interactions?
specificity and affinity
The higher the affinity of the binding, the more likely a specific interaction will take place.
The ____ the affinity of the binding, the ____ likely specific interaction will take place
higher; more
Depending on the context, the cell must always have ways to ____ specific context to make the interaction specific.
regulate
Which protein domain binds to pTyr?
SH2
The measure of the intrinsic strength of the binding interaction.
affinity
The quantitative strength of a binding interaction is defined by the ____.
Kd - dissociation constant
What is the simple binding reaction equation?
A + B <–> AB
For stronger binding, would you predict Kd to be a higher or lower value?
lower
Kd = K(off)/K(on) = ??
[A][B]/[AB]
From [A][B]/[AB], how can Kd be determined?
Kd can be determined by the concentrations of B and AB. A eventually runs out so that is why it plateaus.
What constant relates to the concentration of the substrate needed to make the enzyme work efficiently?
Km (Michaelis’ constant)
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?
Km
How is Kd different from Km?
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.
What does a Kd of 10^-15 M tell us?
The interaction is strong
ex. Avidin-biotin binding
What does a Kd of 10^-7 M tell us?
The interaction is semi-weak
ex. SH2 domain-phosphotyrosine (pTyr) site
What is the thermodynamic relationship of the dissociation constant?
Relative amounts of free components of the complex present at equilibrium are directly related in the free energy of the two states.
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.
lower energy; small amount
What is the term for the idea that the world is attracted to lower energy?
energy minima
What is deltaGº?
intrinsic free energy of the binding
What should deltaGº be in order for protein binding to occur?
It must be negative (-deltaGº)
< 0
How can mutations affect binding?
Small changes in binding energy will cause large difference in Kd. Thus, mutations in binding energy can change Kd.
Which binding would be preffered for protein P?
The binding to B would be more favored because it generates lower energy
The amount of time it will take for the half life of the complex to dissociate.
half-life
What are the assumtions made about a theoretical calculation based on the equation?
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
What is the effect where the presence of multiple binding sites would significantly increase the affinity?
Avidity effect
Ideal affinity and specificity depends on biological function and ligand concentration.
How does the cell control this?
kinases and phosphatases
What could be an advantage to lower affinity?
The protein can adapt to change, more interactions can occur, it is transiet and short-lived
Which of these associations is most likely a heat shock protein?
hsp70/clients
-ATP -> +ATP
Why would the hormone/receptor association have a high affinity and specificty?
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
