Protein sorting Flashcards
What are imported into the nucleus
Histones, transcription factors, DNA/RNA polymerases, RNA splicing factors, DNA repair enzymes
What are exported from nucleus?
mRNA, tRNA, ribosomal subunits, shuttling proteins
What perforates the nuclear envelope?
Nuclear pore complex
What type of arrangement does the NPC have and what else?
Octagonal arrangement
Central aqueous pore
Many individual proteins
Small molecules pass through pores how?
freely by diffusion
How do large molecules get into the nucleus?
Active transport
Nuclear proteins complete synthesis where and pass through pores how?
Complete synthesis on cytosolic ribosomes and pass through pores in fully folded state
How do the right proteins get into the nucleus? what allows them to be targetted to the nucleus
Nuclear localisation signals sequences
What makes up the NLS Sequences
A continuous stretch of amino acids 15-60 residues long
Where are the NLS sequences?
Near the N terminal
OR
A signal patch where there are multiple internal sequences in the unfolded protein which then combine to form a signal patch in the folded protein form
The NLS sequences are recognised by what?
Complementary receptors
Nuclear localisation requires an intact and functional what?
NLS
A sequence rich in what amino acid targets a protein to the nucleus?
Lysine rich
Mutation of Lysine rich sequence to what causes cytoplasmic retention?
Lysine to Threonine
A cargo protein has what on it?
NLS
The nuclear import receptors recognises and binds to what?
NLS on the cargo protein
Nuclear import receptors interact with what to transfer cargos in and out of the nucleus?
Interact with the nuclear pore complex
Describe the binding and dissociation of nuclear import receptors to NPC transports cargo into nucleus
- Cargo with NLS binds NIR
- NIR shutles into the nucleus via the NPC (FG repeats)
- Ran GTP binds to NIR to discharge the cargo
- NIR shuttles out of the nucleus via NPC with the RanGTP bound (FG repeats)
- Ran GTP is hydrolysed to Ran GDP in the cytosol
- NIR is free to shuttle more cargo into the nucleus
Ran GDP is found where and where is Ran GTP
These two forms confer on what?
Ran GDP is in cytosol
Ran GTP is in nucleus
Confers directionallity on transport
Name a way nuclear protein can be regulated
Via NFAT, transcription factor found in the cytoplasm
NLS in NFAT is cryptic but exposed after phosphorylation of amino resides (Ser) by a ca2+ regulated kinase.
Phosphorylation of NLS in NFAT produces what?
a conformational change
Mitochondrial proteins are imported how?
Fully synthesised but unfolded polypeptide chains and chaperones prevent folding.
What are the signal sequences like in mitochondrial translocation?
Amphiphillic, forms an alpha helix with hyophobic, polar and hydrophillic residues on separate side of the helix
Hydrophobic region matches what on the receptor for mitochondria?
Matches hydrophobic groove of receptor
What moves the mitochondrial proteins through both membranes?
Protein translocators
Where are TOM and SAM located?
Outer mitochondrial membrane
Where are TOM, TIM and OXA located?
Inner mitochondrial membrane
The following complexes insert protein into where?
1) TOM
2) SAM
3) TIM
4) OXA
1) Proteins into outer membrane
2) Outer membrane
3) Inner and matrix and inserts mitochondrial derived proteins into inner membrane
4) Inner membrane and matrix
Describe how a protein gets into the mitochondrial matrix?
1) Precursor unfolded protein binds to the import receptor in TOM complex.
2) Passes through the outer mitochondrial membrane and into the inner membrane via the TIM complex .
3) Once in the matrix, signal peptidase cleaves off the signal on the protein and it is folded into a stable structure to produce a mature mitochondrial protein
Mitochondrial import requires what?
Energy ATP
Describe how chaperones and ATP is involved in mitochondrial protein import
Chaperone proteins (Cytoplasmic Hsp70) binds to precursor peptide.
The release requires ATP hydrolysis to push protein through TOM complex.
Import via TIM results in mitochondrial Hsp70 binding
ATP hydrolysis pulls protein through TIM complex
Synthesis of ER targeted proteins begins where and completed when?
Begins on free ribosomes in cytosol but translation is completed after ribosome binds to ER
What directs ER signal sequence to specific receptor in ER membrane
Signal recognition particle
Describe the process of directing a signal polypeptide to the ER membrane
do again
