Molecular Switches

Question 1

What enzyme catalyses glycogen breakdown?

Answer 1

Glycogen Phosphorylase

Question 2

What enzyme catalyses glycogen synthesis?

Answer 2

Glycogen Synthase

Question 3

Allosteric Regulation of Glycogen Phosphorylase

Answer 3

Glycogen Phosphorylase is activated by AMP
AMP is antagonised by ATP and G6P (Glucose-6-Phosphate)
Note: AMP/ATP/G6P bind to an allosteric site (Distinct from the active site)
G6P and ATP are feedback inhibitors of glycogen breakdown (since they are produced from glycogen breakdown)

Question 4

Significance of Phosphorylase activation by AMP

Answer 4

2ADP -> ATP + AMP
Catalysed by adenylate kinase, a reversible reaction with little energy change.
Maintained close to equilibrium in most cells (Keq = 1)

If ADP/ATP rises 2-fold (signifying a fall in cellular energy), AMP/ATP rises by 4-fold

AMP:ATP ratio is thus more sensitive indicator of cellular energy status that ADP:ATP

Because effects of AMP are antagonized by ATP,
phosphorylase is activated by an increasing ratio of AMP:ATP

Question 5

“Homeostatic” versus “Switch” mechanisms:

Answer 5

Allosteric regulation is good for maintaining homeostasis
An allosteric mechanism is switched off by a simple reversal of the on mechanism. However this requires a LARGE change in input to give a large change in output, which is where the switch mechanism comes in.

Switch mechanisms have different on and off pathways (unlike the reversal of the allosteric mechanism)
Switch mechanisms involve covalent modifications of proteins e.g. Phosphorylation

Question 6

Covalent protein modifcations:

Answer 6

Reversible:
- Phosphorylation
- Ubiquitination
- Acetylation
Irreversible:
- ADP-ribosylation: bacteria that cause cholera catalyses ADP-ribosylation

Question 7

Advantages and Disadvantages of Allosteric Regulation

Answer 7

Adv.:
- No enzymes required
- No energy directly consumed by ligand binding
Disadv.:
- Large changes required for large effects

Question 8

Advantages and Disadvantages of Covalent Modifcation Regulation:

Answer 8

Adv.:
- Small changes can give any desired size of effects
Disadv.:
- Must be catalysed by enzymes
- Some ATP consumed (But this is the “price” for precise control)

Question 9

Covalent modification of glycogen phosphorylase:

Answer 9

2 forms of phosphorylase discovered:
- The a form that did not require AMP for activity
- The b form that was allosterically activated by AMP
The a form is phosphorylated while the b form is dephosphorylated form
Ca2+ activates phosphorylase kinase which is responsible for the phosphorylation.
Therefore glycogen breakdown is perfectly synchronised with muscle contraction.

Phosphorylase kinase is itself phosphorylated by cAMP-dependent protein kinase A (PKA), causing an increase in maximal activity and Ca2+-sensitivity.
cAMP is increased in muscle by the hormone adrenaline, released during stress or exercise.
Therefore more rapid and complete conversion of phosphorylase to the AMP-independent a form occurs when the organism is under stress, and adrenaline is circulating

This hormonal mechanism (mediated by cyclic AMP) superimposes on the nervous mechanism (mediated by Ca2+), while both over-ride the allosteric mechanism (mediated by AMP)

Question 10

Why are there multiple tiers of regulation?

Answer 10

For different levels of delay between initiation and response. As small delays as possible required for a quick a response to increase survival chances (Tom and Jerry analogy)

Question 11

The human genome encodes >500 protein kinases

TRUE OR FALSE

Answer 11

TRUE

Question 12

Protein kinase cascades:

Answer 12

One kinases phosphorylating another kinase, causing amplification of the signal.
This signal can converge and diverge to form signalling networks (Like a CPU).

Question 13

A molecular switch without covalent modifications?

Answer 13

G proteins.
Involves a conformational change in a GTP-binding or G- protein.
Basal state: G proteins are inactive with GDP bound
Activate state: G-protein with GTP bound
A signal activates a GTP/GDP exchange factor (GEF) allowing exchange from basal to active state.
The active G-protein binds and activates a downstream effector protein.
In the absence of continued stimulation there is reversal back to the basal state.
GTP hydrolysis is catalysed by the G protein, but promoted by GTPase activator proteins.

Question 14

Irreversible mechanism: Blood Clotting

Answer 14

Uses a cascade of proteinases (clotting factors) that activate each other by proteolysis.
Each clotting factor cleaves an inactive precursor of its downstream target, activating it.
Proteolysis is irreversible and, once used up, clotting factors have to be re-synthesised.