Lecture 12- Allosteric Models

What is Allostery?

• A way of regulating the activity of proteins

• Cooperativity is a type of allostery

• The binding of a molecule at one site on a protein, that’s going to affect change at distinct/diff site on that same protein

• (change will affect activity binding of protein)

• In hemoglobin it affects the binding

Hemoglobin subunits and how they’re subunits go from tightly coupled to bonded with oxygen 

• In the T-state the individual hemoglobin subunits are tightly conformationally coupled to each other. (alpha 1 binds to alpha 2 and Beta 2) (beta 2 binds to Beta 1 and alpha 1 ) (beta 1 binds to alpha 2 and beta 2)

• Oxygen binding induces conformational transition that breaks ionic interactions with neighbors

• Breaking of ionic bonds reduces conformational restraint (further relaxes state) and allows neighbors to adopt high affinity state

• Oxygen can bind to any of the subunits 

What does a sigmoidal binding (ss shaped curve)  curve indicate, explain.

It indicates cooperative binding 

1. Thes first O2 molecules binds weakly to T state subunit

2. T to R transition is easier for second binding 

3. Last molecule binds to a subunit that is already in the R state 

Left on graph is low affinity and right on graph is high affinity

Experimental indicator of Allostery

Cooperative binding ( if you see an SS shaped curve)

what is the hill equation

Shows the fraction of cooperative binding

Tells us if the binding is cooperative or nah (N_H )= slope

• If n_H = 1 No cooperativity 

• If  n_H >1 Positive Cooperativity

• If n_H <1 Negative Cooperativity

For myoglobin: Slope never changes so it is =1

A Hill coefficient of 1 means no cooperative ligand binding is occurring in this particular protein. True for myoglobin

nh cannot exceed n

What is the Monod, Wyman and changeux (MWC) model (concerted Model)

• The MWC model explains cooperativity by saying all subunits switch between low- and high affinity states together, and ligand binding shifts the balance toward the high-affinity state (R). 

1. only 2 conformations of the tetramer exists (T and R)

2. T and R conformations are in equilibrium (can go back and forth)

3. All subunits undergo transition simultaneously 

4. ligand can bind to both T and R states but has  a higher affinity for R

5. Each successive ligand binding increase likelihood of T to R transition (shifts equilibrium so it lies more in the R state as more oxygens bind)

What is the Koshland Nemethy and Klmer (KNF) Sequential model?

• The KNF model says binding happens one site at a time, and each binding slightly changes the shape of the protein — making the next site more likely to bind.

• In reality, hemoglobin behavior fits both models partly/
  it shows concerted-like switching overall (MWC) but also local stepwise changes between subunits (KNF).

1. Ligand can bind both T and R state

2. Ligand binding induces T to R in single subunit

3. Transition in one subunit increases likelihood of transition in adjacent subunit

4. Requires existence of many mixed tetramer conformations

Concerted (MWC) vs. Sequential (KNF) Models

• Sequential works in showing positive cooperativity in hemoglobin since it shows local stepwise changes between subunits (KNF). But usually compatible with negative cooperativity.

• Concerted Model: If we follow 2 parameters below we can come up w/ a predicted way the , hemoglobin will bind and plot data points of fractional binding of hemoglobin at diff conc.

• Controlled by: (1) affinity difference (T vs R), (2) equilibrium position without ligand.

• All subunits switch together → only T or R states, no intermediates.

Pulmanory Respiration

1 of the 2 types of respiration:

We breathe oxygen from lungs and we exhale CO₂ out.

Cellular respiration

1 of the 2 respirations:

Occurs in tissue

Uses oxygen and produces CO₂

How to transport CO₂ safely back to the lungs from tissue?

• CO₂ is not very soluble; it will form bubbles in blood and kill you, bad'

• We must do this rxn:

CO2 + H2O <=> H⁺+ HCO₃ ^-

This reaction gets catalyzed by Carbonic anhydrase to make

HCO₃ ^- which is much more soluble and can be transported back

It also releases more protons, which raises the PH→ Ph in tissues will be slightly more acidic

Carbonic anhydrase

Enzyme that reacts w/ co2 and H2o to produce bicarbonate (HCO3)

Bohr effect

How pH and CO2 concentratration will affect O2 affinity

• High Ph= High O2 affinity→lungs

• Low Ph=Low O2 affinity—→tissues

• High CO2 =Low affinity

• Low CO2= High affinity