1. Chap 10 allosteric 10.1

2. Why regulate enzyme activity?
Regulation allows for efficient use of resources by the cell.
Metabolic pathways rarely stand alone, and usually intersect with numerous other pathways
In enzyme pathways of several steps there are often cataslyzed by key enzymes that are rate-limiting in controlling the flux through the pathway
This integrating expression of the pathway with other metabolic needs of the cell

3. Why regulate enzyme activity?
Regulation allows for efficient use of resources by the cell.
Metabolic pathways rarely stand alone, and usually intersect with numerous other pathways
In enzyme pathways of several steps there are often catalyzed by key enzymes that are rate-limiting in controlling the flux through the pathway
This integrating expression of the pathway with other metabolic needs of the cell

4. Regulating Activity: 4 main strategies
1. Allosteric control. Proteins contain distinct regulatory sites and
multiple functional sites. Binding of regulatory molecules
triggers conformational changes that affect the active sites.
Display cooperativity: small [S] changes - major activity
changes. Information transducers: signal changes activity or
information shared by sites

5. Regulatory Enzymes
Allosteric enzymes undergo conformational changes in response to modulator binding
Subunit interactions in an
allosteric enzyme, and
interactions with inhibitors
and activators

7. Hemoglobin allosteric properties
changes in quaternary structure lead to exposure of binding pocket thus easier access of O2
tertiary structure of low affinity: deoxy-Hb T (taut) state
tertiary structure of high affinity: HbO2 R (relax) state

8. allosteric kinetics
Allosteric enzymes DO NOT obey Michaelis Menten kinetics!
At low substrate concentration, the enzyme is inactive thus in the in the inactive conformation.
As the substrate concentration is increased, substrate binds to enzyme and triggers a conformation change to the active conformation of the enzyme.
After the first substrate is bound, the second and subsequent substrates all bind more readily “cooperativity”

9. allosteric kinetics
At low substrate concentration: the enzyme is in inactive conformation
As the substrate concentration is increased: substrate binds to enzyme and subsequent substrates all bind more readily
Allosteric enzymes “S-shaped curve” indicates cooperative binding of substrate. The result of the change to the active shape is a steep increase in both substrate binding, and as a result, in reaction rate over a narrow range of substrate concentration.
When all active sites on the allosteric enzyme are occupied with substrate, a plateau is reached.

10. allosteric kinetics
Allosteric enzymes can be regulated between very low and very high reaction rates with only small changes in substrate concentration.
Allosteric enzymes are used by cells to regulate metabolic pathways where the concentration of cellular substrates fluctuate over narrow concentration ranges.

11. aspartate transcarbamoylase
ATC-ase

12. Two views of the regulatory enzyme ATCase T R
This allosteric regulatory enzyme has two stacked catalytic clusters, each with three catalytic polypeptide chains (in shades of blue and purple) and three regulatory clusters, each with two regulatory peptide chains (in red and yellow).
Modulator binding produces changes in enzyme conformation and activity

13. Structure of ATCase - side view
Side view of quaternary structure

14. R and T states in equilibrium

15. ATCase does not obey MM kinetics
Substrate binding to one active site converts enzyme to R state
increasing their activity: active sites show cooperativity

16. Basis of sigmoidal curve
R & T states equivalent to 2 enzymes with different K0.5s

23. On the following plot, N represents the curve for an allosteric enzyme with no allosteric activators or inhibitors added. If an allosteric activator was added, which curve would one obtain?