Question

Concerted opening. Suppose that a channel obeys the concerted allosteric model (MWC model, p. 200). The binding of ligand to the \(R\) state (the open form) is 20 times as tight as that to the T state (the closed form). In the absence of ligand, the ratio of closed to open channels is \(10^{5}\). If the channel is a tetramer, what is the fraction of open channels when \(1,2,3,\) and 4 ligands are bound?

Short answer

Use the partition function to calculate open channel fraction for each ligand count bound. The fraction increases with more ligands bound due to higher R state affinity.

Step-by-step solution

Understanding the Model

The MWC model describes how the binding of a ligand to a protein can change the protein's conformation. In this model, the protein can exist in two states: the Relaxed (R) state, which binds ligands tightly, and the Tense (T) state, which binds ligands less tightly.

Defining Parameters

Given that the binding affinity of ligand to R is 20 times stronger than to T, we define the binding constants as follows: \( K_R \) for R state and \( K_T = 20 K_R \) for T state. The ratio of closed to open channels in the absence of ligand is \( L_0 = 10^5 \), where \( L_0 = \frac{[T]}{[R]} \).

Calculating the Partition Function

Calculate the partition function \( Z \) for the tetramer in terms of ligand binding sites: \[ Z = (1 + K_T[L])^4 + L_0(1 + K_R[L])^4 \] where '[ ]' denotes concentration, and '4' comes from the fact there are 4 sites on the tetramer.

Calculating Probability for Each Ligand Binding

For a given number \( n \) of bound ligands, calculate the fraction of open forms:- For no ligands bound (n=0): \[ P_0 = \frac{(1)^4 + 10^5 (1)^4}{Z} \]- For 1 ligand bound (n=1): \[ P_1 = \frac{\binom{4}{1} K_T[L] + 10^5 \binom{4}{1} K_R[L]}{Z} \]- For 2 ligands bound (n=2): \[ P_2 = \frac{\binom{4}{2} (K_T[L])^2 + 10^5 \binom{4}{2} (K_R[L])^2}{Z} \]- For 3 ligands bound (n=3): \[ P_3 = \frac{\binom{4}{3} (K_T[L])^3 + 10^5 \binom{4}{3} (K_R[L])^3}{Z} \]- For 4 ligands bound (n=4): \[ P_4 = \frac{\binom{4}{4} (K_T[L])^4 + 10^5 \binom{4}{4} (K_R[L])^4}{Z} \]where \( \binom{4}{n} \) is the binomial coefficient.

Evaluating Fractions Numerically for Bound Ligands

Plug in \( L = 1 \) (standard concentration) and numerically solve the probabilities using the previously derived equations for each \( n \) from 1 to 4. Simplify the expressions using the ratio of binding constants and the defined \( L_0 \).

Interpreting the Results

From the calculated probabilities, determine the fraction of open channels for 0, 1, 2, 3, and 4 bound ligands. The highest fractions will be associated with states where more ligands are bound, favoring the R state.

Key concepts

MWC Model

The Monod-Wyman-Changeux model, often called the MWC model, describes how proteins transition between different states in response to ligand binding. It's particularly useful for understanding cooperative binding, where ligand binding at one site affects the binding properties at another site on the same protein. This model posits that proteins like ion channels can exist in two conformations:

• The Relaxed state (R), which has higher affinity for the ligand.
• The Tense state (T), which has lower affinity for the ligand.

The MWC model assumes the transition between these states is concerted, meaning that the entire protein shifts from one state to the other all at once, rather than individual subunits changing independently. This synchronized behavior is like a switch being turned on or off, providing a simplified framework to model complex biological systems.

Ligand Binding

Ligand binding is a fundamental concept where small molecules (ligands) interact with binding sites on proteins, much like keys fitting into locks. In the MWC model, the ligand binding affinity in the R state is described as being much higher compared to the T state. Specifically:

• The R state binds ligands 20 times more tightly than the T state in our given scenario.
• Binding affinity is described by binding constants, denoted by \( K_R \) for the R state and \( K_T \) for the T state, where \( K_T = 20 K_R \).

The interaction of a ligand with the binding site causes conformational changes, shifting the equilibrium towards the state that binds the ligand more strongly, thereby stabilizing that form. These transitions and their accompanying changes in binding affinity are central to understanding how proteins like channels and enzymes are regulated by ligands.

R and T States

The R (Relaxed) and T (Tense) states are the heart of the MWC model, representing the two primary conformations a protein can adopt. These states are fundamentally linked to the protein's binding affinities and biological functions.

• R State: The high-affinity state which binds ligands very tightly. It is often associated with increased biological activity.
• T State: The low-affinity state which binds ligands less effectively. It is typically more stable in the absence of a ligand.

In the absence of ligands, proteins are predominantly in the T state, which favors stability. However, as ligands bind, they push the equilibrium towards the R state, reflecting the protein's shift to a more active/high-affinity conformation. This binary switch between R and T states is a useful mechanism to regulate activity and efficiency in complex biological systems.

Tetramer

A tetramer is a complex formed by four subunits bonded together, and it's a common structural form in proteins, including ion channels. In the context of the MWC model and ligand binding:

• Each subunit can bind a ligand, contributing to the overall protein's conformational state using cooperative binding dynamics.
• The tetramer structure implies there are four possible binding sites for ligands, significantly influencing the protein's behavior under the concerted allosteric model.

The interaction of a tetramer with ligands provides insights into cooperative binding—where the binding of one ligand affects the binding properties at other sites—and the overall functional state of the protein. This structure makes it possible for proteins to exhibit allosteric behavior, whereby the binding at one site can enhance or inhibit binding at another site, crucial for the fine-tuning of biological activities.