Lecture 15- Protein degradation by the ubiquitin-proteasome system Flashcards
Ubiquitin basic structure
beta sheet around an alpha helix, flexible c-terminal tail- allows recognition by a lot of proteins
why is protein degradation importany
preventing cytotoxicity (removing damaged or aggregating proteins), fine-tuning the levels of a protein within a cell
where is Ub attached
lysine side chains, c-terminus attached to the gamma-amine by an isopeptide linkage
enzymes involved in Ubation- steps
Ub attaches to E1- ATP dependent
E1 transfers Ub to a Cys on E2
E2 collaborates with E3, catalyses isopeptide bond formation
variations in each enzyme type
2 E1s, 40ish E2s, varying numbers of E3s- 500 human 1200 A. thaliana
what is the ubiquitin code
repetitive Ubation of a protein, which can have distinct consequences for a cell
how many lysines in Ub
7
examples of different Ub chain topologies and their function
K63 (on 63rd lysine)- non-proteolytic functions
K48- proteolysis
what does PolyUb do
targets the protein towards the 26s proteasome, a ‘degron’
what is the 26S proteasome
major eukaryotic proteasome, responsible for protein degradation in the cytosol and nucleus
2 functional parts of the proteasome
barrel-like 20S core particle, contains active sites
19S regulatory particle, acts as a cap at one/both ends
what is the benefit of having separate subunits
allows for compartmentalisation, and high levels of control
rough structure of 20S core
heptameric rings, beta subunits are proteolytically active and cut at different points into peptides of average length 7-8aas
how is access to the chamber regulated
opening and closing of axial pores via changes in the N terminal tails of the alpha subunits
regulatory subunit composition
Rpn (non-ATPase) and Rpt (triple A ATPase) subunits
how do substrates move into the core through the RP
loops projecting from ATPases push the substrate through and unfold it
examples of Rpn subunits and their roles
Rpn10- Ub receptor, provides a ‘platform’ to load peptides
Rpn 1- scaffold protein
6 subunits forming a ‘lid’, acts as a stabilising anchor
how many ‘stages’ can the proteasome exist in
4- where s1 has a closed core gate, and s4 has an open gate (subunits are sligned)- s2 and 3 are intermediates
what changes the state
substrate binding can lead to conformational shifts
prerequisites for proper folding into the proteasome
polyubiquitin tagging and a disordered region
what is ERAD
ER associated protein degradation
4 main steps of ERAD
recognition
dislocation and ubiquitination
targeting
proteosomal degradation
example of ERAD pathways
ERAD-L (at the Lumen)
ERAD-M (at the Membrane)
-named based on location of the degron
function of Cdc48
retrotranslocation motor- pulls proteins out of the ER to deliver them to the 26SP
