intracellular transport

·      Why can’t proteins be transported via channels or transporters like an ion?

-       Because they are too large and complex

·      What is a signal sequence/sorting signal? What are some examples of signal sequences?

-       Bound by proteins or transmembrane proteins in organelle membranes to allow passes into the nucleus or mitochondria

·      True or false: all proteins not translated on mitochondrial or chloroplast ribosomes are at least partially translated in the cytosol

true

·      True or false: free ribosomes and membrane bound ribosomes are similar in structure, but encoded by different genes in the genome

false

Signal recognition particle:

-       ribonucleoprotein that recognizes and binds to the signal sequence emerging from ribosome translating a protein destined for ER. Stops further protein elongation

srp receptor

-       docking site for srp-ribosome complex, binding srp. Triggers the release of the signal sequence and the ribosome from srp allowing polypeptide to enter channel

Protein translocator

-       protein-conducting channel that opens when docking, allows the polypeptide chain to pass through the ER membrane or becomes integrated into the membrane.

·      Think of an example of a protein that would need to be transported into the nucleus of a eukaryotic cell.  What path would this protein need to take from beginning translation to its final destination? What sorting signals would need to be present?  

-       Translation starts in the cytosol: ribosomes in the cytosol begin translating histone mrna

-       No ER signal: since its for the nucleus, it lacks ER signal, so translation continues on free ribosomes

-       NLS (nuclear localization signal) recognition: NLS is exposed and recognized by import receptors

-       Nuclear pore complex docking: histone complex moves the NPC

-       translocation complex goes through NPC in an energy dependent process

-       release (nucleus): inside the nucleus, histone is released to perform function

·      Think of an example of a protein that would need to be transported to the plasma membrane of a eukaryotic cell.  What path would this protein need to take from beginning translation to its final destination? What sorting signals would need to be present? 

-       Rough endoplasmic reticulum (RER): translation begins on free ribosomes in the cytoplasm. N-terminal signal directs the ribosome-mrna to RER membrane, protein goes into the ER or inserted into membrane

-       Endoplasmic reticulum: proteins fold, assemble, and sometimes get glycosylated (sugar added).

-       Golgi apparatus: proteins move to ER to golgi in vesicles. Further processing, modification and sorting occur

-       Vesicular transport protein destined for the plamsa membrane go off the golgi in transport vesicles

-       Plasma membrane: vesicles move and fuse with plasma membrane, putting protein in correct orientation

o   What signal sequences would you expect to be present in a histone protein?

-       Nuclear localization signal

o   Consider an enzyme like pyruvate dehydrogenase.  What signal sequences would you expect to be present in that enzyme? What path would it take from the beginning of translation to its final destination?

-       Mitochondrial targeting sequence

-       Path starts at translation on free ribosomes in cytosol

-       Mitochondrial targeting sequence directs protein to the surface of mitochondrion after translation finishes

-       Protein is recognized by mitochondrial membrane

-       Protein is translocated across membrane to mitochondrial matrix

-       Signal sequence is taken off by signal inside matrix

o   Consider a voltage gated ion channel. What signal sequences would you expect to be present in that enzyme? What path would it take from the beginning of translation to its final destination?

-       Start and stop-transfer sequences

-       Translation starts on free ribosome in cytosol

-       Hydrophobic sequence is recognized by signal recognition particle which stops

·      Consider examples where a protein could contain conflicting signal sequences (like an ER signal sequence and an NLS).  Which would “win” and why?

-       The ER would win because it’s the first to emerge from the ribosome during translation. Its recognized by the signal recognition particle which stops translation and directs the ribosome/mrna complex to the ER membrane, and the protein enters the ER as its being synthesized

·      What is the endomembrane system? Which cell organelles are part of the endomembrane system?

-       Lipids and proteins move through it in transport vesicles surrounded by a phospholipid bilayer

·      Mitochondria are membrane bound but are not considered to be part of the endomembrane system.  Why?

-       Because they aren’t derived from the er/golgi, don’t go protein/lipid transport, and have their own dna/double membrane

·      Where are proteins destined for the endomembrane system translated?

-       Free ribosomes in the cytosol

·      Describe the important proteins and steps in ribosomal docking

-       Small ribosomal subunit binds to mrna near 5’ cap

-       The complex scans the mrna until it finds start codon

-       Initiator trna dos into the p-site of small subunit

-       Large ribosomal subunit joins complex, forming complete ribosome with A-cite open

·      What is the difference between a soluble protein and a transmembrane protein? What sequences must a transmembrane protein have that a soluble protein does not?

-       Soluble proteins are hydrophilic and float freely in aqueous environments, while transmembrane proteins are embedded in cell membranes which require hydrophobic regions to interact with lipids and hydrophilic parts to face cytosol

·      What is a vesicle and what do vesicles do in the cell?

-       Bud off of one membrane and fuse with another, transferring their phospholipids and their cargo

·      What happens in the golgi apparatus (very generally) and how does this relate to the final destination of proteins?

-       Receives proteins from the er, modifies, sorts ,and packages them into vesicles and adding specific labels that direct them to their final destinations