Post-translational Processing Of Proteins

Question 1

What are the 2 types of post-translational modification?

Answer 1

Proteolytic cleavage - breaking peptide bonds to remove part of protein
Chemical modification - addition of functional groups to amino acid residues

Question 2

What proteins are synthesised on free ribosomes?

Answer 2

Protein destined for cytosol or post-translational import into organelles

Question 3

What proteins are synthesised by ribosomes on RER?

Answer 3

Proteins destined for membrane or secretory pathway via co-translational insertion

Question 4

What is required for protein sorting?

Answer 4

A signal, intrinsic to the protein
A receptor that recognises signal and which directs it to correct membrane
Translocation machinery
Energy to transfer protein to new place

Question 5

What proteins are involved in targeting for secretion?

Answer 5

Extracellular proteins
Membrane proteins
Vesicular proteins (lysosomes, endosomes)

Question 6

What are the 2 types of secretion from cells?

Answer 6

Constitutive - constantly secreted (e.g. albumin)

Regulated - endocrine, exocrine and neurocrine cells

Question 7

What is a signal sequence?

Answer 7

N-terminal aa sequence
5-30 amino acids in length
Central region rich in hydrophobic aa residues
Able to form alpha-helix

Question 8

What does the pre in preproalbumin mean?

Answer 8

Signal sequence which is removed during processing

Question 9

What is the signal recognition particle (SRP)?

Answer 9

It’s the receptor needed to bind the signal peptide on proteins destined for the ER
Composed of 6 proteins and short piece of RNA
Recognises signal peptide and ribosome

Question 10

Describe the synthesis of secretory proteins and their translocation across the ER membrane

Answer 10

Ribosome binds to mRNA, with signal sequence outside ribosome
SRP recognises signal sequence and binds to the ribosome and stops protein synthesis
SRP receptor forms part of the translocation machinery
SRP binds to receptor which is GTP dependent
Translocon opens and the SRP is released so protein synthesis can start again
Signal peptidase in the membrane cleaves the signal sequence and protein is translated into the ER
Ribosome then detaches from mRNA

Question 11

What are the functions of the ER?

Answer 11

Glycosylation
Formation of S-S bonds
Proper folding of proteins

Question 12

What is N-linked glycosylation?

Answer 12

Sugars are added on asparagine side chain (reaction involves amino group)

Question 13

Why is glycosylation important?

Answer 13

Correct protein folding
Protein stability
Facilitates interactions with other molecules
Deficiencies in N-linked glycosylation lead to severe inherited human disease

Question 14

What does protein disulphide isomrase do?

Answer 14

Role in formation of disulphide bonds in ER lumen

Question 15

What happens if there are folding problems?

Answer 15

Protein may be trapped in mis-folded conformation
Protein contains mutation resulting in mis-folding
Protein may be incorrectly associated with other sub-units

Question 16

What do ERchaperone proteins do?

Answer 16

Attempt to correct the problems in protein folding
It retains unfolded proteins in ER
Acts as sensors to monitor extent of protein mis-folding

Question 17

What happens in mis-folding cannot be corrected?

Answer 17

Protein may be returned to cytosol for degradation

Proteins may accumulate to toxic levels in ER resulting in disease

Question 18

What is the basic unit of collagen fibres?

Answer 18

Tropocollagen

• 3 polypeptides (alpha chains)
• glycine in every 3rd position
• characteristic triple helix

Question 19

What are the features of the collagen triple helix?

Answer 19

Non-extensible
Non-compressible
High tensile strength

Question 20

Why is glycine the amino acid in the collagen structure?

Answer 20

Glycine is the only amino acid with a side chain small enough to fit in middle of helix

Question 21

How are the alpha chains held together in a collagen fibre?

Answer 21

H bonds between alpha chains stabilise

Question 22

What are the steps of synthesis and modification of collagen in ER?

Answer 22

1. Synthesis and entry of chain into lumen of RER
2. Cleavage of signal peptide
3. Hydroxylation of selected proline and lysine residues
4. Addition of N-linked oligosaccharides
5. Addition of galactose to hydroxylysine residues
6. Chain alignment, formation of disulphide bonds between alpha chains
7. Formation of triple-helical procollagen from C- to N- terminus

Question 23

What is prolyl hydroxylase?

Answer 23

Associated with PDI (protein disulphide isomerase) in ER
Requires vit C and Fe2+ ions for activity
Allows increased H bonding to stabilise triple helix
Scurvy due to weak tropocollagen triple helices

Question 24

What happens at the C and N terminals of the collagen fibre?

Answer 24

150 extra N-terminal amino acids do not form triple helix
250 extra C-terminal amino acids do not form triple helix
Called propeptides

Question 25

What are the steps of procollagen secretion?

Answer 25

1. Completion of O-linked oligosaccharide chains by addition of glucose
2. It is then transported in a vesicle
3. Exocytosis occurs
4. Removal of N and C terminal propeptides

Question 26

What enzyme breaks peptide bonds to remove terminal peptides?

Answer 26

Procollagen peptidase
Procollagen -> tropocollagen
This occurs extracellularly

Question 27

Describe the steps of formation of collagen fibres

Answer 27

1. Removal of N and C terminal propeptides
2. Lateral association of collagen molecules followed by covalent cross linking
3. Aggregation of fibrils