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 Presentation

Allostery

2017-10-27

You measure MM curves for a variety of enzymes (solid curves). You add molecule $A$ to each solution and remeasure its MM curve (dashed line). For each curve: What changed ($K_{M}$ or $V_{max}$)? Is this a competitive or noncompetitive inhibitor?
$K_{M}$, competitive
$V_{max}$, noncompetitive
$V_{max}$, activator

Competitive inhibitor

• Inhibitor competes with substrate for same site
• This raises $K_{M}$: lower apparent affinity for substrate because it has to compete
• No effect on $V_{max}$. If you add enough substrate, you swamp out competitor
• Inhibitor is chemically similar to the substrate
• Example: Ethanol competes with methanol for alcohol dehydrogenase, lowering the rate of formaldehyde production.

Noncompetitive inhibitor

• Binds distant from the active site, altering active site to turn off activity
• This leads to a drop in $V_{max}$. (lower $[E]_{T}$: Less of the enzyme is in the active form.)
• No effect on $K_{M}$. What active enzyme is around has exact same affinity for substrate
• The inhibitor need not have any chemical similarity to the substrate

Example noncompetitive inhibitor

Conceptual goals

• Understand that binding at one site in a protein can alter activity at another site
• This "allostery" arises because the "allosteric effector" interacts with one conformation, but not the other.
• Understand how this applies to hemoglobin (BPG).

Skill goals

• Determine how binding at on site affects activity at the other
• Predict the effects of adding allosteric effectors to a system of equilibria.

Hemoglobin transports $O_{2}$ from the lungs to tissues

stephaniefuturedoc

Hemoglobin is a tetramer four proteins

Credit: Janet Iwasa (Utah)
Credit: Janet Iwasa (Utah)
Credit: Janet Iwasa (Utah)

$E_{active} + I \rightleftarrows E_{inactive} + I \rightleftarrows E_{inactive} \cdot I$

$[E]_{active} = [E]_{T}\theta_{active}$

$\theta_{active} = \frac{[E_{active}]}{[E_{active}] + [E_{inactive}] + [E_{inactive}\cdot I]}$

What are the ingredients you would need for allostery?

Ingredients:

• Two different binding sites recognizing different things
• Equilibrium between two (or more) protein shapes
• Different "activities" (functions, properties, etc.) of each shape
• Binding to one shape, but not the others

Summary

Allostery is when binding at one site in a protein alters activity at another site

• Two different binding sites recognizing different things
• Equilibrium between two (or more) protein shapes
• Different "activities" (functions, properties, etc.) of each shape
• Binding to one shape, but not the others

Noncompetitive inhibitors

BPG allosterically regulates $O_{2}$ binding in hemoglobin

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