Lecture 12 - Ubiquitin II Flashcards
How do different Polyubiquitin Linkage types encode different signals?
(3 Points)
- Polyubiquitin Chains possess different topologies (3D Shapes) depending on the linkages involved
- These different shapes have different interaction landscapes, allowing interaction with different proteins
- Different linkage types also possess different conformational flexibility, and may adopt more filamentous or compact structures
Define Ubiquitin Receptors (Ub-R)
Family of Proteins which recognise ubiquitin modifications via non-covalent interactions, decoding the “Ubiquitin Code” to regulate various processes
Define Ubiquitin Code
Structurally Unique Ubiquitin Linkage types recruit a distinct set of effector proteins (Ub-R) which influence fate of modified protein
By What Two ways can a Lys-48 linked chain be recognised by the 26S proteasome?
- S5a subunit of 19S Cap is a Ub-R, which can recognise K48-chain directly
- S2a subunit of 19S Cap recognises K48-Chain indirectly via interaction with Ub-R Hhr23a
Give an Example of the role of K63-linked chains in signal transduction
- Many receptors (e.g., IL-1R, TNFR) utilise TRAF6 for signal transduction, with activation of receptor leading to its polyubiquitination (K63-linked)
- TAB2 (Ub-R) recognises K63-linked chains, and exists in complex with TAK1 (kinase)
- TAB2 binding to pUb chain results in conformational change, which induces a conformational change in TAK1 that activates it
(i) How do Ub-R’s recognise ubiquitin modifications (Give Examples)?
(ii) How strong are these interactions?
(i) Ub-R’s utilise short protein sequences known as UBDs to recognise ubiquitin via non-covalent interactions (e.g., NZF, UIM, UBA)
(ii) Low Affinity interactions, hence most Ub-Rs contain UBDs in tandem
What regions of a Ubiquitin monomer can be recognised by a UBD?
- Hydrophobic Surface (centred around I44)
- Acidic Surface (centred around D58)
- C-terminal tail (Gly76) - if free
How can polyubiquitin chain linkage impact the ability of UBDs to bind to specific surfaces (Provide one example and an exception)?
Chains with different linkages adopt different topologies, which may occlude a particular UBD binding site
Example - in K48-linked Diubiquitin chains, the two I44 hydrophobic surfaces may interact non-covalently, making it more difficult/impossible for a UBD to recognise
Exception - Hhr23a UBA domain interacts with two adjacent hydrophobic surfaces to provide avidity
* UBA domain - 3 helix bundle which has 2 Hydrophobic surfaces
How can UBDs bind in tandem to increase affinity?
(2 Methods)
- Single Ub-R possessing multiple UBDs (e.g., USP5)
- Monomeric Ub-Rs may oligomerise together (Avidity)
Why is the Spacing of UBDs in tandem important?
- Different Ubiquitin Linkages produce different distances between interaction surfaces, therefore UBDs must be spaced correctly to allow high affinity interactions (i.e. 3D Molecular Ruler)
- Tandem UBDs are optimally positioned to interact with unique conformations of polyubiquitin chain (e.g., Lowest Energy Structure)
Describe USP5 in terms of:
(i) What it is?
(ii) Function
(iii) Structure
(iv) Affinity
(i) Deubiquitinating Enzyme
(ii) Degrade Unanchored polyubiquitin chains, which can be bound by RIG-I, activating its kinase activity and leading to innate immune responses
(iii) Consists of:
* Two UBA domains - recognise I44 Hydrophobic Surface
* One ZnF-UBP - recognises terminal G76 of free ubiquitins
(iv) Multivalent interactions generate relatively high affinity (nM)
Give Two Examples of Ubiquitin-mediated Processes, and the Linkages Involved
- Proteosomal Degradation - typically K48-Linked, although all but K63-Linked can signal degradation
- Trafficking (e.g., ESCRT Pathway) - involves monoubiquitin, with UBDs often increasing in affinity along the pathway (i.e. lowest first)
Give Two Examples of Diseases associated with dysregulation of Ubiquitin System
- Familial Parkinson’s (E3 Ligase Parkin)
- Cancer (E3 Ligase BRCA1)
