what is the lysosome’s main function
recycling macromolecules and organelles
what is the lysosome’s environment like
- it has a low pH maintained by proton pumps
- it is filled with digestive enzymes
what’s the signal sequence for lysosomal proteins
mannose-6-phosphate
how is the mannose-6-phosphate signal formed
1) proteins that will become lysosomal enzymes receive N-linked oligosaccharides containing mannose
2) then, in the cis-golgi, N-acetylglucosamine phosphotransferase adds a phosphate group to mannose, creating mannose-6-phosphate
what enzyme adds a phosphate onto the mannose oligosaccharide
N-acetylglucosamine phosphotransferase
how are proteins sent to lysosomes via the golgi
1) after the m6p signal is translated, it will bind to an m6p receptor in the golgi
2) this binding will recruit adapter proteins, then clathrin coated vesicles will form
3) through GTP hydrolysis the clathrin coat will shed once the vesicle buds off
4) the vesicles will fuse into endosomes
5) eventually the phosphate signal will be cleaved off, allowing the protein to dissociate from the m6p receptor
6) from endosomes they will be transported to fuse with the lysosomes
what prevents lysosomal enzymes from being active too early
the enzymes work at low pHs, so the compartments that the enzymes are carried in are at higher pHs
what do you call a protein prior vs after to its activation
- prior: pro-protein
- after: active-protein
what is a lysosomal storage disease
- neurodegenerative brain disease
- accumulation of lipids and protein waste in neurons
- incurable childhood disease
what is mutated in severe lysosomal storage diseases
- the N-acetylglucosamine phosphotransferase enzyme
- so lysosomal enzymes won’t be targetted to the lysosome, they will be secreted
what’s a potential treatment for lysosomal storage diseases
enzyme replacement therapy: when diseased cells were grown in vitro, they took up the missing enzyme (N-acetylglucosamine phosphotransferase) via endocytosis
what’s a challenge with enzyme replacement therapy
it’s difficult to make the actual enzyme
what is endocytosis
process of taking up (swallowing) fluid, particles or molecules from external medium
what’s a type of endocytosis cells undergo
receptor-mediated endocytosis
describe the process of receptor-mediated endocytosis
1) ligand, like m6p, binds to receptor on the extracellular cell surface, which has an internalisation signal
2) receptor recruits AP2
3) AP2 recruits clathrin on the intracellular cell surface
4) clathrin accumulates until it causes an inwards curvature of the cell
5) dynamin wraps around the neck of the vesicle and uses GTP hydrolysis to pinch it off
6) after entering the cell, the clathrin coat disassembles and AP2 detaches
what is LDL
- low density lipoprotein
- the “bad fat”
- packages of hundreds of lipid molecules in a
large water-soluble carrier
what do we want to happen with LDL in our bloodstream
be taken in by cells and broken down by lysosomes
how is LDL taken into cells
1) LDL in the bloodstream binds to LDL receptors which have an internalisatino sequence
2) this triggers receptor-mediated endocytosis
3) the LDL will be delivered to the lysosome
4) the low pH level will cause LDL to dissociate from its receptor and enter the lysosome
5) in the lysosome, the LDL will be broken down into amino acids, fatty acids and cholestrol
is the receptor-mediated endocytosis of LDL a quick or slow process
quick: ~6 minutes
what is base editing
- changes one base in the gene
- this can be done in vivo
how can base editing be used to treat high LDL
1) there’s a gene that codes for an enzyme enabling endocytosis of LDL receptors
2) endocytosis of LDL receptors = LDL in the blood can’t be broken down
3) by mutating these genes, the receptors won’t be endocytosed and so LDL can continue to be intaken and broken down
what are the 2 ways proteins can be released
- regulated secretion
- constitutive secretion
how does regulated secretion work
1) pre-pro-protein made in ER
2) signal sequence removed to make pro-protein
3) pro-protein processed in golgi then packaged into secretory vesicles
4) inside vesicles, pro-protein is cleaved to make mature protein
5) vesicles wait near the plasma membrane - release only happens when a signal arrives!
how does constitutive secretion work
1) pre-pro-protein made in ER
2) signal sequence removed to make pro-protein
3) pro-protein processed in golgi
4) in secretory vesicle, pro-protein cleaved to make mature protein
5) vesicles immediately fuse with the plasma membrane so the protein is constantly secreted into the blood - there is no storage step!
