Session 8 - Protein Function and Regulation Flashcards
Name 5 short term regulatory mechanisms
1) Different enzyme forms - isoenzymes
2) Change in enzyme conformation - allosteric regulation
3) Reversible covalent modification - phosphorylation
4) Proteolytic activation
5) Controlling the amount of enzyme present - gene expression
Explain how isoenzymes can be used to regulate functions and give an example
Enzymes that catalyse the same reaction but are different proteins. They have:
- different levels of activity
- different regulatory properties
Example: Hexokinase and Glucokinase
(These can be synthesised from different genes or differently spliced from the same gene)
Explain how allosteric regulation works
Exist in two forms: T state = low affinity
R state = high affinity
These states are brought about by the binding of activators/inhibitors:
Activators - increase the proportion of enzyme in the R state
Inhibitors in the proportion of enzyme in the T state
Example: Phosphofructokinase-1
Explain how reversible covalent modification can act as a form of regulation
Protein kinases - add phosphates to an -OH group via the use of ATP
Protein phosphatases - work in the opposite direction
- This bonding gives the compound lots of free energy so that it is able to undergo further reactions
How does proteolytic activation work as a form of regulation?
- These are inactive precursor molecules - Zymogen or Proenzymes
- These can then have the “pro” segment removed via hydrolysis of a peptide bond and they will be activated
Example Digestive enzymes, clotting factors
Outline the regulation of the clotting process
1) Inactive zymogens present at low concentrations
2) Proteolytic activation
3) Amplification of initial signal by cascade mechanism
4) Clustering of clotting factors at site of damage
5) Feedback activation by thrombin ensures continuation of clotting
6) Termination of clotting by multiple mechanisms
7) Clot breakdown controlled by proteolytic activation
What is the role of thrombin (Prothrombin) in blood clotting?
1) Prothrombin is activated by factor X to form thrombin
2) Thrombin cuts off fibrinopeptides on fibrinogen (fibrinopeptides prevent fibrinogen coming together) to form fibrin
3) Fibrin monomers assemble via non-covalent interactions (soft clot)
4) Covalent cross-linking occurs between Lys and Gln catalysed by transglutaminase (Factor XIII)
Thrombin also acts as a positive feedback on factors V,VIII,XI,XIII to further drive the clotting
How is the clotting process stopped and a clot then broken down?
Stopping - 1) Dilution of clotting factors and removal by liver
2) Digestion by proteases
3) Binding of inhibitors
Breaking - Plasmin degrades fibrin to fragments it is formed from plasminogen and activated by t-PA and streptokinase
What are the main differences between haemoglobin and myoglobin?
Haemoglobin - In blood, Transport O2 around body in blood, 4 polypeptide chains, 4 haem groups per molecule.
Myoglobin - In muscle, Short term storage of O2, 1 Peptide chain, 1 haem group per molecule
Why does the affinity of 02 to myoglobin stay constant yet in haemoglobin it changes?
Haemoglobin is constituted of 4 haem groups each binding to their own 02 as one 02 binds there is a slight conformational change in there shape and as such this affects the other haem groups bonding to O2. Myoglobin however only has one sub unit and as such the conformational change has no effect on the binging of other molecules to it
Name some allosteric regulators of 02 binding
1) 2.,3 BPG - lowers affinity - stabilises T state
2) CO2 and H+ - lowers affinity - stabilises T state
What effect does CO have on haemoglobin?
- CO binds 250x more readily than 02
- It blocks further O2 binding once bound
- Stabilises R state in unaffected subunits - preventing dissociation at tissues
Explain what happens in sickle cell disease
1) mutation of glutamate to Valine on Beta Globin (HbS)
2) Valine lies on surface in T state (deoxygenated) and causes haemoglobin molecules to stick to one another
3) Rigid rods form increasing cells likely hood to lyse leading to anaemia. And also block microvasculature which causes pain
What are the functions of the endoplasmic reticulum?
- Insertion of proteins into membrane
- Specific proteolytic cleavage
- Glycosylation
- Formation of S-S bonds
- Proper folding of proteins
- Assembly of multi-subunit proteins
- Hydroxylation of lysine and proline residues
What are the functions of the Golgi Apparatus?
- Movement of proteins to the cis-Golgi through budding
- Further protein modifications
- Release through trans-Golgi to membrane/organelles
