Describe the structure of the 26S proteasome.
2.5 MDa
structure:
19S RP (regulatory particle), consisting of base (AAA+ ATPases, protein unfolding) and lid (substrate recognition and deubiquitination)
20S CP (catalytic particle), consisting of αββα subunits (proteolysis)
19S RP
The parts of the RP must be aligned to form a continuous channel to obtain the active proteasome.
Which steps are necessary to degrade a protein to single amino acids? Which complexes/enzymes are involved?
proteasome: 10-14 aa
tripeptidyl peptidase: 2-4 aa
aminopeptidase: single amino acids
How are ATP and GTP used in the cell?
ATP is a general energy currency
protein synthesis
DNA synthesis
muscle movement
transport of macromolecules
synthesis of cellular components
phosphorylations
GTP is more involved in signal transduction and protein synthesis
How is protein homeostasis maintained in eukaryotes?
key systems: ubiquitin-proteasome system, protein unfolded response, chaperone system
balance between protein synthesis and degradation/control; when one is out of control: problem
homeostasis: maintain balance between protein synthesis and quality control
Name two major protein degradation pathways in eukaryotes.
proteasome/ubiquitin system
autophagy system
Which cellular systems monitor protein folding?
chaperone systems
Name abundant AAA ATPase family proteins.
Abundant AAA ATPase family proteins:
proteasome
NSF (snare disassembly)
Orc1 (DNA replication)
Name the known functions of AAA+ ATPase family proteins.
diverse roles in the cell, for example involved in:
DNA replication
protein degradation
membrane fusion
peroxysome biogenesis
protein disaggregation/refolding
DNA recombination/repair
transcription
…
Which amino acids are included in the pore‐loop that interacts with extended polypeptides?
Phe, Val, generally rather unpolar amino acids
Outline the mechanism by which AAA+ ATPases unfold substrates.
tunnel with (mostly) hydrophobic handles that pull the protein through a hole
ATP hydrolysis!
1 ATP à 5-8 residues
initiation (“Einfädeln”) is the difficult part
What is the number of ATPase subunits required to constitute a proteasome ATPase motor?
six
Name three peptidase activities of the proteasome.
β1: caspase-like protease – cleaves after Asp, Glu
β2: trypsine-like protease – cleaves after basic residues (Arg, Lys)
β3: chemotrypsine-like protease – cleaves after Phe, Tyr
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