BIOLOGY 2B03 Lecture Notes - Lecture 10: Translocon, Sec61, Secretion
6 Jun 2018
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Protein Targeting: Co-translational Targeting
Rough Endoplasmic Reticulum (RER)
1) bound by a single membrane •
2) no genome - everything coded in nuclear genome •
3) site of protein synthesis and modification •
4) first destination for secreted proteins •
Are Proteins Targeted While Translation is Active
Hypothesis: if transport occurs during translation, then proteins will exit •
ribosome and go directly into a microsome
Microsomes: created by disrupting ER membranes via homogenization ◦
pieces of membrane assemble into spherical structures that mimic normal ◦
ER behaviour
Method: microsome membranes treated with detergent to release proteins inside •
released because added protease digests proteins ◦
when microsome membranes are intact and protease is added, newly synthesized proteins are protected in ◦
interior of microsome
Observation: newly synthesized proteins are protected from proteases within microsome •
Conclusion: newly synthesized proteins move directly into microsome •
Must Translocation and Translation Occur Together
Hypothesis: if translation is completed, transport cannot occur •
Method: proteins synthesized in vitro with microsomes - mature proteins found on inside •
proteins also synthesized without microsomes and when translation is complete, the proteins without ◦
ribosomes added to solution with microsomes - these proteins enable to enter microsomes
Observation: transport only occurs when translation occurs in presence of microsomes •
Conclusion: import MUST occur co-translationally •
Co-Translational Transport - 5 Characteristics again!
1) Signal sequence on transported protein •
2) Receptors for signal Sequences •
3) Translocation channels •
4) Energy requirement - GTP hydrolysis for ER •
5) A way of targeting proteins to the different locations within an organelle •
Rule 1 - Is There a Peptide Signal Sequence
16-30 AA N-terminal signal: •
short positively-charged domain ◦
central hydrophobic domain ◦
polar domain ◦
ER signal sequence directs active ribosome to ER membrane •
Rule 2 - Is There a Signal Receptor
Signal Recognition Particle (SRP), made of 6 proteins - binds to: •
ER signal sequence on protein ◦
large ribosomal subunit ◦
SRP receptor ◦
signal must be at N-terminus for co-translational transport - if at C-terminus, translation ceases before SRP •
binding
transport won't happen without ribosome - SRP receptor has hydrophobic binding groove for ER signal sequence •
