Proteins - Post translational Modification Flashcards
Where are proteins made on free ribosomes in the cytosol targeted for?
Cytosol
Organelles - e.g. nucleus, mitochondria, peroxisomes
Where are proteins made on RER bound ribosomes targeted for?
For ER, Secretion/Lysosomes/Plasma membrane
Protein targeting: Which out of ER, Lysosome, Mitochondria, Nucleus, Peroxisome
1) Doesn’t require direct energy
2) Has a C terminal signal
3) Doesn’t require receptor
4) Is kept unfolded on entry to organelle
1) ER - indirect by H+ pump to keep lysosome pH5
2) ER - KDEL is C terminal, Peroxisome SKL is C terminal.
3) Nucleus - importin through pores
4) Mitochondria - chaperones keep unfolded
What is constitutive vs regulated secretion? What 3 cell types tend to do regulated secretion?
Constitutive constant - collagen, albumin
Regulated - at specific times e.g. insulin
Endocrine/exocrine/neurocrine
Where are signal sequences on secretory proteins e.g. collagen/albumin, how many amino acids, are they hydrophillic/phobic, and what do they form to help get across membrane?
N terminus
5-30 amino acids
Central region hydrophobic
Can form alpha helix
What does the ‘pre’ part denote in preproalbumin etc
The signal sequence
What are the 5 basic stages to synthesis of secretory proteins and their translocation across the ER membrane?
1) Signal - N terminus
2) SRP - binds signal and ribosome - halts translation
3) SRPreceptor translocon binds all of above
4) Peptide enters - signal sequence cleaved by signal peptidase
5) Translation continues - fully folded proteins
What are the basic steps of preproinsulin processing in ER/Golgi? (4)
1) In ER- signal peptidases cleaves signal sequence
2) Folding of proinsulin with 3 x cysteine-cysteine disulphide bonds
3) Endopeptidases cleave B chain
4) To golgi - mature insulin 2 peptides joined with disulphide bonds packages in zymogen granules for secretion
What are the differences in membrane protein biosynthesis compared to secreted after the signal sequence has been cleaved by endopeptidases (3)?
1) Membrane proteins have a stop transfer sequence which halts protein in membrane
2) Moves out of translocon = transmembrane domain
3) Translation continues and more transmembrane domains are made if needed
What 7 functions of ER in protein procesing?
Proteolytic cleavage Proper folding Insertion into membrane Assembly of multi subunits Glycosylation Hydroxylation of Lys Pro Dilsuphide bonds
3 reasons glycosylation of proteins is so important - what disease can occur if there is a deficiency in N-Linked glycosylation
1) Correct protein folding
2) Protein stability
3) Facilitates interactions with other molecules
Congenital disorders of glyosylation - very severe
Where is the sugar added to in N-Linked glycosylation?
Asparagine side chain
Which enzyme plays a role in formation of disulphide bonds in ER?
Protein Disulphide Isomerase
What do chaperone proteins in ER do if proteins are misfolded? Give examples of chaperones that do this
Can actively fold/restrict folding to allow for misfiled proteins to be refolded correctly.
E.g. BiP - a Heat Shock Protein 70, Calreticulin, Calnexin
What if misfolding cannot be corrected (2)
1) Back to cytosol for degradation
2) May accumulate in ER - toxic
Can a single mutation lead to aggregation of misfiled proteins in ER?
Yes
Apart from recruiting chaperones to help refold misfolded proteins, what other thing can the cell do to help prevent this happening?
Stop translation
How and where does O linked glycosylation occur?
Golgi
Adds sugar to OH group of Ser Thr
What kind of glycoslyation do you see in EC matrix/mucus secretions? What do these two things have in common relevant to this kind of glycosylation?
O linked - important in Proteoglycans.
What are the 4 ‘Threes’ of tropocollagen structure? Which are the most common AA to be in the X and Y positions?
triple helix
300nm rod
Gly XY (Gly every 3rd AA)
3 polypeptide chains
Commonly proline and hydroxyproline in XY positions
Is tropocollagen non extensible?
Yes-
Is tropocollagen non compressible?
Yes
Where does Gly sit in the triple helix? Why?
In the middle - small
Where are the H+ bonds formed between the helices in tropocollagen? Which attribute of collagen do the H+ bonds give rise to? What other bonds mean collagen is very structurally stable?
Proline gets hydroxylated post translation - forms H+ bonds
High tensile strength
Cross link covalent bonds between collagen molecules. Disulphide bonds between cysteine residues.
