The languages of signal transduction- post translational modification

Question 1

What are post translational modifications

Answer 1

1. Reversible covalent modifications that are used extensively by cells to regulate protein function.

Question 2

What are 4 forms PTMs can take place as

Answer 2

1. Addition of small chemical groups (phosphorylation, acetylation etc.)
2. Addition of large molecules: sugars, proteins (e.g. ubiquitination) and lipids
3. Cleavage of proteins/proteolysis.
4. Isomerisation

Question 3

What are properties of PTMs

Answer 3

1. Change is rapid, specific, flexible, and tightly regulated.

Question 4

How does PTM enables proteins to exist in many different states

Answer 4

1. PTM allows a huge variety of modifications, and there are a large number of different sites on a protein that can be modified.
2. This greatly increases the number of potential molecular states of a protein.
3. Provides versatility: each single genetically encoded protein is actually many different proteins in the cell.
4. (~20,000 protein-encoding genes in humans).

Question 5

How does PTM save energy

Answer 5

1. In the absence of PTM, major changes in the protein makeup of a cell would require new gene transcription and protein synthesis.
2. PTM saves energy!
3. they are centrally important for virtually all signal transduction mechanisms.
4. New protein synthesis is energetically costly to the cell. Moreover, the synthesis of new protein is a relatively slow process.

Question 6

Describe conformational changes due to PTM

Answer 6

1. In conformation of the polypeptide backbone.
2. Or in the intramolecular arrangement of the different folded domains of the protein.
3. Diverse effects on activity:
4. Stimulating or inhibiting catalytic activity.
5. Altering the ability to bind to other proteins.
6. Enabling further post-translational modifications by unmasking or burying potential modification sites.

Question 7

Describe altered protein-protein binding due to PTM

Answer 7

1. PTM can create or destroy a binding site due to effects on charge distribution, hydrogen-bonding possibilities and shape of binding surface
2. PTM can also change interaction of proteins with other cellular components, such as nucleic acids or membrane lipids
3. Changes in protein interaction often have secondary effects – such as changes in subcellular localisation, or further PTMs.

Question 8

Show how PTMs enable formation of signalling complexes with the example: Insulin receptor substrate (IRS1):

Answer 8

1. Adaptor protein containing a PTB (phosphotyrosine binding) domain and a PH (pleckstrin homology) domain.
2. Each interaction on its own is too weak to cause significant recruitment of IRS1, and therefore both phosphotyrosine and phosphoinositide motifs must be present to form a signaling complex.
3. First, the activated receptor phosphorylates itself on tyrosines, and one of the phosphotyrosines then recruits a docking protein called insulin receptor substrate-1 (IRS1) via a PTB domain of IRS1;
4. the PH domain of IRS1 also binds to specific phosphoinositides on the inner surface of the plasma membrane.
5. Then, the activated receptor phosphorylates IRS1 on tyrosines, and one of these phosphotyrosines binds the SH2 domain of the adaptor protein Grb2.
6. Next, Grb2 uses one of its two SH3 domains to bind to a proline-rich region of a protein called Sos, which relays the signal downstream by acting as a GEF to activate a monomeric GTPase called Ras.
7. Sos also binds to phosphoinositides in the plasma membrane via its PH domain.
8. Grb2 uses its other SH3 domain to bind to a proline-rich sequence in a scaffold protein.
9. The scaffold protein binds several other signalling proteins, and the other phosphorylated tyrosines on IRS1 recruit additional signalling proteins that have SH2 domains.

Question 9

Describe how PTM affects Subcellular localization

Answer 9

1. PTM can alter dynamics of shuttling of proteins between different sub-cellular compartments (termed protein trafficking).
2. E.g. lipid modification directs many proteins to stably interact with cellular membranes.
3. E.g. transmembrane proteins release an active fragment that diffuses away and exert its effects elsewhere in cell

Question 10

Describe how PTM affects Proteolytic stability

Answer 10

1. Alters expression level of a protein.
2. Some phosphorylation events can target a protein for proteolytic degradation, whereas other modifications can specifically stabilize a protein.
3. Phosphorylation is often coupled to ubiquitylation, resulting in proteolytic degradation.

Question 11

Common post-translational modifications (PTMs)

Answer 11

1. Addition of simple functional group
2. Addition of large molecules
3. Proteolysis and isomerisation

Question 12

What are some simple functional groups that can be added

Answer 12

1. Phosphorylation- Phosphorylation in prokaryotes where it is important component of bacterial chemosensing
2. Methylation
3. Acetylation
4. Hydroxylation

Question 13

What are some common amino acids which are modified

Answer 13

1. Serine
2. Threonine
3. Tyrosine
4. Proline- hydroxylation
5. Lysine- Methylation found in histones
6. Arginine- Methylation and acetylation found in histones
7. Glutamate
8. Histidine

Question 14

What are some large group addition s

Answer 14

1. Glycosylation
2. Ubiquitination
3. Addition of lipids

Question 15

Where is hydroxylation of proline found

Answer 15

1. Hydroxylation found in exm proteins- collagens

2. Scurvy caused by insufficient hydroxylation of prolines

Question 16

What are PTMs caused by

Answer 16

1. PTMs are catalysed by specific enzymes and, in many cases, are reversed through the action of other enzymes (‘writer’ / ‘eraser’ enzyme pairs).