regulation of protein degradation Flashcards
pathways of protein degradation (3)
lysosomes
extracellular proteases
ubiquitin-proteasome pathway
reasons why cells degrade their own proteins (4)
1: physiological removal of rate-limiting enzymes and regulatory proteins
2: quality control of misfolded proteins
3: usage of proteins as energy source (primarily in skeletal muscle during catabolic states)
4: antigen processing by immune system (extracellular antigens processed by endosome/lysosome pathway and intercellular by ubiquitin-proteasome pathway)
ways proteases are classified (3)
by:
active site nucleophile
- serine
- cysteine (lysosmal proteins)
- threonine (proteasome proteases)
- water (metalloproteases and acidic proteases)
substrate specificity for:
- positively charged AA (eg trypsin)
- negatively charged AA
- hydrophobic AA (eg chymotrypsin)
cleavage location
- aminopeptidase - can only start at N terminus
- endopeptidases - can cleave protein anywhere
- carboxypeptidase - can only start at C terminus
ways cells regulate protein degradation (4)
1: natural protease inhibitors (eg serpins)
2: inactive enzyme precursors (zymogens) - enzyme must be activated like prohormones, etc.
3: compartmentalization (lysosomes)
4: specific death signals (ubiquitin)
note: proteolytic degradation of peptide bonds is energetically favorable - need ATP to regulate the protein degradation process
lysosomal pathway of protein degradation
1: primary lysosomes formed from Golgi apparatus
2: these fuse with endosomes (have membrane proteins or cells that bind to membrane proteins outside cell like LDL) or autophagosomes (bulk canabalism - takes up large amount of cytoplasm and proteins by engulfing it with membrane - stimulated in states of starvation) to make secondary lysosomes
3: lysosomal proteases break down contents
regulation of lysosomal pathway
proteases (pepsins in lysosomes) regulated by:
1: compartmentalization - put membrane around pepsins
2: requirement for acidic conditions (provided by ATP-dependent lysosomal proton pump - pepsins only active at low pH)
3: specific cathepsin inhibitors (called cystatins) - inhibit any lysosomal proteins that leak out
pathway also regulated by hormonal signals during catabolic states
cathepsin K deficiency - pycnodysostosis
cathepsin K involved in regulation of lysosomal degradation of proteins
deficiency results in genetic disorder of bone development
extracellular protease pathway of protein degradation
- required in which processes?
synthesized by cells and secreted into bloodstream
1: tissue remodeling (matrix metalloproteases and elastase)
2: complement cascade (immune response)
3: clotting (thrombin cascade)
4: fibrinolysis (aka plasmin cascade; breaking down of the clot)
5: digestion (pancreatic proteases)
serpins
natural serine protease inhibitors regulate extracellular proteases genetic diseases associated with serpin deficiency: 1: alpha1-antitrypsin deficiency 2: C1INH deficiency 3: antithrombin deficiency
ubiquitination steps
for selective protein degradation for proteins in cytoplasm, nucleus and ER
1: ubiquitin-activating enzyme (E1) activates carboxyl group of ubiquitin - E1’s active site is cysteine - results in reactive thioester - uses ATP, generates AMP + 2 Pi
2: activated ubiquitin is transferred to carrier proteins (E2)
3: E2 binds to E3 to make ubiquitin ligase complex (E1 there too)
4: degradation signal on target protein binds to E3 (E3 has binding pockets specific for certain proteins, or for an aspect of misfolded proteins such as a hydrophobic domain)
5: complex facilitates transfer of ubiquitin to e-amino groups on lysine residues in substrate proteins => isopeptide bond - energy neutral way so don’t need more ATP
6: the E1 that had the ubiquitin leaves and is replaced by another E1 that has an ubiquitin, which can then be attached to the target protein, allowing for polyubiquitination
7: proteasome then degrades ubiquinated peptide into small peptides and AA
note: cells have different combinations of E2 and E3 and different combinations recognize different protein substrates
proteasome steps
multienzyme complex
1: entry lid has receptor proteins that bind to ubiquitin-tagged proteins - only binds polyubiquitinated ones
2: deubiquitinating enzymes (isopeptidases) cleave isopeptide bonds and ATP-dependent enzymes (both types of chaperone proteins) unfold the target proteins
3: proteases within cylindrical core have differing substrate specificities - hydrolyze proteins in central channel of proteasome
note: all proteases threonine proteases (no other threonine proteases known)
diseases involving ubiquitin-proteasome pathway
cervical cancer
parkinson’s
multiple myeloma (plasma cell tumor) treated with proteasome inhibitors
dynamic state of cellular proteins
always turning over
example: crystalin, life of 80 years, as age turnover slows, why need lens replaced
hemoglobin, life of 120 days
sperm cells - exzyme ornithinie decarboxylase enzyme has life of 11 minutes
anabolic state
body builders on steroids
catabolic state
diabetes, starvation
physiological removal of rate-limiting enzymes and regulatory proteins
cyclin degraded to allow movement out of G2 into mitosis
inflammation regulated this way too
protein quality control
when proteins not folded properly
if have hg point mutation, unstable so is degraded
can have normal DNA sequence too - post translational modifications occur incorrectly or damage to protein after its been folded - by reactive O2 species (esp. due to smoking)
can have transcription errors (30% of transcripts have errors)
or folded improperly (like if chaperones aren’t available)
clinical correlation: cystic fibrosis - chloride channel is mutated so that cell mistakes it for being folded wrong or a mistake and it gets degraded
another example: alcoholic liver damage - proteins in liver cells denatured due to long term exposure to alcohol
use of proteins as energy source
if malnourished, use for energy to get AA for gluconeogenesis
protein degradation as antigen processing by the immune system
necessary for both MHC1 and MCH2
alpha1-antitrypsin deficiency
clinical correlation for extracellular protease protein degradation
smoker with emphysema and liver failure
many neutrophils in lungs and protein deposits in liver
genetic, recessive
1/1000 people of european descent
alpha1-antitrypsin inhibits elastase secreted by neutrophils at site of inflammation - allows itself to be target for elastase and cleaved by it - intermediate clings to elastase so inhibits it (suicide substrate)
if deficiency, less effective substrate for elastase - smoking makes it worse because more inflammation
get misfolded alpha1-antitrypsin protein aggregating in liver
curable with liver transplant
anaphylactic shock
recurrent tongue swelling and shortness of breath
can be spontaneous - not due to allergy
genetic lack of C1 inhibitor - get massive compliment activation, angioedema
antithrombin deficiency
clinical correlation for protease deficiency
pregnant woman develops blood clot in leg and lung after miscarriage
thrombosis in leg that breaks off and lodges in lung
predispose to it by antithrombin deficiency - thrombin converts fibrinogen to fibrin to create clot
biochemistry of ubiquitin attachment
ubiquitin attaches to internal lysine group in protein
new form of chemistry - isopeptide bond
cervical cancer
HPV that causes cervical cancer has E6 viral proteins - degrades p53
velcade
new treatment for cancer
proteasome inhibitor that can kill multiple myeloma cells - more sensitive to drug than other cells
