Proteolysis - Latency and Activation Flashcards
What are pre-peptides?
Signal peptides responsible in translocation and translation through membranes
Secretory proteins which are synthesised with signal peptides at the N-terminus which is removed after sorting by proteolysis
What are Pro-peptides?
Folding intramolecular chaperones.
These must be removed for activity to occur.
How does latency relate to the secretory pathway?
Most bioactive polypeptides must be inactive whilst in the secretory pathway and then become activated when they are released.
(proteins synthesised in an inactive form)
What does the Trans-Golgi network do?
Sorting of protein and membrane components to constitutive and regulated pathways and to lysosomes.
Cleavage of some pro-proteins also occurs.
What is a secretagogues?
A substance that causes another substance to be secreted.
Where does the constitutive pathways deliver its components to?
The plasma membrane
Where does the regulated pathway deliver its components to?
Forma pool of contents awaiting a secretagogues for release into the cells
What is essential for activation of a protein?
Pro-peptide cleavage.
What do cleavage sites contain?
At least 2 basic residues
What removes the 2 basic residues in cleavage sites after cleavage?
Carboxypeptidase
What are PCSKs?
Proprotein Convertases Subtilisin/Kexin (prohormone converting enzymes)
They are serine proteases and so will cleave at these sites on pro-peptides.
How were PCSKs found?
Found by homology with Kex 2 from yeast.
What was the first mammalian PCSK found and how many are there in total?
First found was Furin (PCSK3)
Currently 9 known mammalian PCSKs
What are the general structure of PCSK genes?
Signal peptide
Prosegment (synthesised in inactive forms)
catalytic domain
P-domain
What is required for PCSK activation?
Synthesised with a prosegment which requires autocatalytic cleavage to activate the PCSK.
What is the role of the P-domain in the PCSK gene?
Regulates the activity of the catalytic domain
Reacts to pH and ion concentration changes.
What do PCSK specifically cleave?
Cleave mostly C-terminal to arg
What amino acids are recognised by PCSKs?
Arg/Lys
What dictates PCSK specificity?
Dictated by a sequence 2-5 amino acids N-terminal to the bond cleaved.
What is the preference cleavage site for PCSK1,2,3?
PCSK1 -RR (arg, arg)
PCSK2 - KR (Lys, Arg)
PCSK3 - RXKR (Arg, Any AA, Lys, Arg)
What PCSKs are found in the secretory pathway?
PCSK1 and 2
What PCSKs are found in the constitutive pathway?
PCSK 3, 5, 6, 9
What are the transmembrane PCSKs?
PCSK 3, 5, 7
What is special about PCSK3?
Recycles through the plasma membrane and endosomal compartments
What are the important roles for PCSKs?
Pro-peptide cleavage
Protein maturation - Insulin
Receptor recycling - LDL
Exploitation by pathogens - toxin activation
What is the role of PCSK in pro-peptide cleavage?
Provide control for mature protein production
Allow tissue-specific expression of peptide hormones
from common precursors.
What is the role of PCSK in insulin maturation?
Synthesised as preproinsulin (pre is cleaved after synthesis)
Proinsulin is inactive precursor.
Processing by PCSK2 and then PCSK1 to yield mature insulin within the regulated secretory vesicle
What is the role of PCSK9 in LDL receptor recycling?
Role in LDL cholesterol metabolism
Promotes LDL receptor degradation in the lysosome
Prevents LDLR recycling to the plasma membrane
Reduces LDLR number at the plasma membrane
Reduces clearance of LDL from circulation
What can PCSK9 be used to treat?
Hypercholesterolaemia
What do viruses exploit PCSKs for?
For the maturation of envelope glycoproteins
potential therapeutic target
What is the role of PCSK3 in toxin activation in bacteria and plants?
These species contain PCSK3 cleavage sites.
A chain acts as a pro-peptide targeting the toxin to the membrane
PCSK3 cleaves the A chain and liberates the B chain.
B chain can cross the secretory pathway membrane and inactivate ribosomes
What cleavage events occur at the cell surface?
Matrix metalloproteinases act on the ECM
Secretases liberate active domains from membrane bound proteins
What are the roles matrix metalloproteinases?
Cell surface cleavage
Role in cancer invasion
What are the important roles in secretases?
Cell surface activation of proteins
Role in Alzheimer’s
How are matrix metalloproteinases activated?
Activated by Zinc
What is the purpose of the cystine switch in the pro domain of the matrix metalloproteinases?
During the folding, the cystine inhibits the catalytic activity of the metalloproteinases and therefore requires cleavage of the pro form to release the cystine residue and activate the protein.
What is the importance of the modification of the structure of the ECM by matrix metalloproteinases?
Proliferation
Apoptosis
Morphogenesis
What are the roles of metalloproteinases in prostate cancer?
A prostate tumour cell synthesises pro-MMPs which can then be activated when zin is present. This means the MMP can degrade the ECM and allow for cell migration and metastasis
What are the secretases?
Alpha - redundant group of ADAM9, 10, 17
Beta - BACE aspartyl protease
Gamma - presenilins
What are the roles of alpha secretases?
2 domain proteases
Active at or close to the cell surface
Responsible for a wide range of sheddase activities
Give an example of a molecule that ADAM proteases activates?
EGF activation
What is APP?
Amyloid precursor protein
What is Alzheimer’s disease characterised by?
Formation of amyloid plaques in the brain by secretases.
What is the role of APP?
Expressed in neuronal and extra-neuronal tissues and have roles in adhesion, migration and gene expression
How are amyloid plaques formed?
Beta and gamma secretases produce Abeta plaques so inhibition of secretases cause be a therapeutic use of Alzheimer’s disease.
