L17 Post-translational Protein Modification

Question 1

What is a proprotein?

Answer 1

An inactive peptide or protein that needs post-translational modifications to be activated

Question 2

What is post-translational modification?

Answer 2

A proprotein is folded and cut to form its active shape.

e.g. when insulin is being formed, ribosomes feed the growing peptide chain (preproinsulin) into the ER where the signal protein is cleaved by signal peptidase, forming proinsulin.

This is then folded, disulphide bonds are formed and then the C chain is removed to form mature and active insulin.

Question 3

Why do cells need the capability to modify proteins?

Answer 3

Protein synthesis is limited in it’s abilities - only a relatively limited number of polypeptide chains can be made.

Covalent modifications can expand a protein’s potential by altering its spatial structure and biological activity. Allows for millions of proteins to be made.

Question 4

Is post-translational modification reversible?

Answer 4

Some PTMs are reversible (e.g. acetylation, phosphorylation and methylation). This allows rapid dynamic regulation of a protein’s activity.

Some PTMs are irreversible.

Question 5

Post-translational modification:

What is proteolytic cleavage?

Is it reversible?

Answer 5

Cleavage of a protein at a peptide bond.

One or several amino acids could be removed from N-terminus of a protein, or protein peptide bond could be cleaved in the internal part of the protein.

Proteolytic cleavage is irreversible.

Question 6

Post-translational modification:

What is proline isomerisation?

Answer 6

A change in proline residue spatial conformation (transition between cis- and trans- conformations of peptide bonds involving proline).

Can seriously affect protein structure adopted.

Question 7

Which enzyme group catalyses phosphorylation?

Answer 7

Kinases

Question 8

Which enzyme group catalyses dephosphorylation?

Answer 8

Phosphatases

Question 9

What is controlled by cyclins and their cyclin-dependent kinases?

Answer 9

The cell cycle

Question 10

Fill in the gaps:
Pyruvate dehydrogenase is regulated by __1__/__2__ by a protein kinase that is __3__ by high [NADH]:[NAD+] and [acetylCoA]:[CoA], and __4__ by pyruvate.

Answer 10

1) phosphorylation
2) dephosphorylation
3) activated
4) inactivated

Question 11

Give an example of autophosphrylation

Answer 11

EGF pathway - growth factor binds to its receptor and causes the receptor to change shape. The change in shape allows two receptors to come together and phosphorylate one another.

Question 12

When a tyrosine residue is phosphorylated, what does this promote?

Answer 12

Protein:protein signalling interactions

Question 13

Complete the sentence:

You can use phospho-specific antibodies and 2-dimension phosphopeptide mapping to __?__.

Answer 13

…detect phosphorylated proteins.

Question 14

What is protein acetylation?

Answer 14

Protein acetylation is the process in which acetyl group, donated by acetyl Coenzyme A, is transferred to an acceptor amino acid, lysine, in protein.

Question 15

How is protein acetylation catalysed?

Answer 15

The reaction is catalyzed by a Protein AcetylTransferase (PAT).

Process of a protein deacetylation is catalyzed by a Protein Deacetylase (PDAC).

Question 16

What proteins are the most characteristic targets of protein acetylation?

Answer 16

Histones.

The histone PATs and PDACs are called histone acetyltransferases (HATs) and histone deacetylases (HDACs).

But they often have non-histone substrates too.

Allows unravelling of DNA for gene transcription.

Question 17

What is protein methylation?

Answer 17

Protein methylation is the process in which methyl group, donated by S-adenosylmethionine, is transferred to an acceptor protein.

Question 18

How is protein methylation catalysed?

Answer 18

The reaction is catalyzed by a protein methyltransferase. Process of a protein demethylation is catalyzed by a protein demethylase.

Question 19

Which 2 amino acids are usually targets for methylation?

Answer 19

Arginine and lysine

Question 20

Are all protein methylation modifications reversible?

Answer 20

No

Question 21

What is citrullination?

Answer 21

Citrullination involves altering the chemical nature of arginine, converting it to citrulline (a new amino acid) by removing the N group.

Question 22

What is a problem associated with citrullination?

Answer 22

Immune system attacks citrullinated proteins - citrullination is implicated as a cause in auto-immune diseases and arthritis.

Question 23

What is glycosylation?

Answer 23

Addition of mono- and oligo- saccharides to a protein to form a glycoprotein.

Significantly affects protein folding, conformation, distribution, stability and activity.

Question 24

True or false: most cellular proteins are glycosylated.

Answer 24

True - glycosylation plays numerous biological roles:

stability, trafficking, and recognition

Question 25

Where does N-linked and O-linked glycosylation occur?

Answer 25

In the ER and Golgi apparatus

Question 26

What is O-linked glycosylation?

Answer 26

Sugar added one in a time in Golgi, or in cytoplasm.

The sugar is added usually to hydroxyl- group of serine or threonine. In some proteins hydroxy-
lysine or hydroxyproline are glycosylated

Question 27

What is N-linked glycosylation?

Answer 27

Polysaccharide is added as a 14
sugar unit to asparagine residue of the newly synthesised polypeptide in the ER.

Further processing takes place in the Golgi apparatus (removal and addition of sugars)

Question 28

What is the main difference betwen O-linked and N-linked glycosylation?

Answer 28

In contrast to N-linked glycoproteins, O-linked glycoproteins are formed by addition of one sugar at a time, usually consist of a few residues.

Golgi – for secreted proteins

Cytoplasm – for cellular proteins

Question 29

What is ubiquitination?

Answer 29

The last glycine in Ubiquitin is attached to lysine in proteins

Ubiquitin is a small protein containing 76 amino acids.

Question 30

What enzymes are involved in ubiquitination?

Answer 30

Ubiquitination requires three types of enzyme:

• ubiquitin-activating enzymes (E1)
• ubiquitin conjugating enzymes (E2)
• ubiquitin ligase enzymes (E3)

Deubiquitinating enzymes (DUBs) remove it

Question 31

What does attachment of a polyubiquitin chain mean for a protein?

Answer 31

It is marked for degradation in a proteasome.

Proteasomes are large protein complexes inside all eukaryotes and archaea (as well as some bacteria) whose main function is to degrade unneeded or damaged proteins.

Question 32

What is the purpose of lipidation?

Answer 32

To increase the membrane affinity of proteins.

Question 33

Each type of lipidation gives proteins distinct membrane affinities, but all types increase hydrophobicity of a protein and therefore its affinity for membranes.

What are the 4 types of lipidation?

Answer 33

C-terminal glycosyl phosphatidylinositol (GPI) anchor

N-terminal myristoylation

S-myristoylation

S-prenylation

Question 34

Which PTMs are generally irreversible?

Answer 34

Amino acid modification

Cleavage