Protein Targeting: Nucleus Flashcards
What is the size of the nuclear pore complex?
125 million Da
What is the diameter of the nuclear pore complex?
150 - 200 nm
What is the Nuclear Pore Complex composed of?
- Octagonal ring structure embedded in the nuclear envelope
- 8 filaments that extend 100nm in nucleoplasm and are joined at their ends to form the nuclear basket
- Cytoplasmic filaments extending into the cytoplasm
What size of ions, molecules and some proteins does the nuclear pore complex allow to diffuse freely?
Up to 60 kDa
What do GTPase Activating Proteins (GAP) do to the G-protein?
Switches it to a GDP bound state (OFF)
What do Guanine Nucleotide Exchange Factors (GEFs) do to the G-protein?
Switches to a GTP bound state (ON)
What are the TWO characteristics of Nuclear Localisation Signals (NLS)?
Neccessary & Sufficient
Which protein recognises NLS?
Importins
What forms the Nuclear Import Receptor?
A heterodimer of Importin α and Importin β
What is the role of Importin α?
Binds to basic NLS sequence
What is the role of Importin β?
Mediates transport through the nuclear pore complex
What part of the nuclear pore complex does Importin β and the cargo (cargo complex) interact with?
FG-nucleoporins
* Phe (F) and Gly (G) repeats
Diffusion is non-directional so how are nuclear proteins selectively imported to the nucleus where their concentration is higher than in the cytosol?
- Cargo complex is rapidly and selectively disassembled in nucleus
- Once disassembled from importin, the cargo protein cannot diffuse out of nucleus
Which enzyme regulates the assembly and disassembly of the cargo complex?
Ran GTPase
What is the role of Ran GTPase in the NUCLEUS?
- GEF keeps Ran in its GTP bound state
- Binding of Ran-GTP to Importin causes dissociation of cargo complex
- Ran-GTP bound to Importin is exported to Cytosol
What is the role of Ran GTPase in the CYTOSOL?
- GAP causes Ran to hydolyse GTP and convert to GDP bound state
- Ran-GDP dissociates from Importin
- Importin binds to another cargo protein
- Ran-GDP binds to NTF2 (nuclear transport factor 2) and returns to nucleus
Where is Ran-GEF located?
Bound to chromatin so remains in the nucleus
Where is Ran-GAP located?
Associated with the cytoplasmic filaments of the nuclear pore
Which amino acid are Nuclear Export Signals (NES) rich in?
Leucine
What is the Nuclear Export process?
- Exportin-1 forms a complex with Ran-GTP and binds to NES of cargo protein
- All three proteins diffuse through the NPC
- At the exit of the pore, Ran-GTP is converted to Ran-GDP and cargo complex is disassociated
- Cargo protein is released into the cytosol
- Ran and Exportin-1 return separately to the nucleus
What is the major difference between nuclear import and nuclear export in terms of Ran?
Ran is part of the cargo complex in nuclear EXPORT
What are TWO examples of regulated nuclear transport?
- T-Cell Activation
- Steroid Hormone Receptor activity
