The individual compartments have different functions due to the unique profile of proteins

in these compartments.

How do the proteins reach their correct destinations?

Where is the information for destination stored?How is this information read?What are the trafficking patterns?

Cellular addresses of proteins reside in the primary and secondary structure

Cellular sentinels recognize these addresses by direct interaction.

1. Label nascent proteins with radioactive a.a.2. Homogenize vesicles3. Digest +/- detergent4. SDS PAGE

Co-translation

Prove it!

Secretory proteins move from the roughER lumen through the Golgi and then to the cell surface.

1. Transport vesicles2. Cisternal progression3. TGN4. Regulated and constitutive secretion5. Lysozomes

Many of the molecules involved in this process were obtained either via genetic or biochemical approaches.

But, before that, the pathway had to be discovered and carefully characterized.

George Palade got the prize for doing this! see Classic Expt. 17.1

Pulse-chase using acinar cells.

Read Classic Experiment 17.1

TS invertase mutants do not secrete invertase at the nonpermissive temperature.Pulse chase experiments were used to identify were secretion was blocked. Analysis of such mutants revealed 5 classes. Cloned genes indicated molecular components of the secretory pathway

Translocation of secretoryproteins across the ER

Signal Sequence

• Microsomes required• Signal sequence cleaved• Signal peptidase in lumen• SS= charge/hydrophob/charged• ca. 70 amino acids from add-in• A few are different (alpha factor)• This expt. enabled discovery of factors such as the SRP

40

30

(1) 300-bp RNA plus (6) subunits

SRP relieves stall

Chem.-modifiedlysyl tRNA

Orientation of membrane proteins is established during co-translation.