Protein Trafficking

1. Site of protein synthesis
   1. Synthesized by ribosomes

1. Processing of proteins
   1. Proteins Secreted in vesicles

1. Budding Mutation
   1. accumulation in ER
      1. Distended ER
2. Vesicle fusion mutation
   1. vesicles accumulate in cell

1. Incubation
   1. radioactive amino acid (Pulse)
      1. One group is immediately fixed
2. 2nd incubation
   1. non-radioactive amino acids (chase)
      1. Cells fixed at various time points post pulse-chase
3. "labeled" proteins closer and closer to PM over time
4. Movement is towards apical side

1. Transfection
   1. Uses live organism
   2. allows to watch prot. from start to finish
2. GFP DNA isolated
   1. fused with target section of DNA
      1. Stop codon removed
      2. two protein blueprints fused
         1. One protein coded by two "Blueprints" fused
            1. Originally from RNA
3. Example of use:
   1. study temp. sensitive muts.
      1. allows observation of location of specific protein

1. Homogenization
   1. Microsomes
      1. Formed from the homogenate
      2. formed by endomembrane system
         1. Different microsomes
            1. Different enzymes
      3. Still contain transmembrane proteins
2. Centrifugation
   1. density gradient
      1. TM proteins isolated
         1. Production of anti-bodies against protein
            1. endomembrane markers
               1. allow to tack protein trafficking
   2. Separates ER from Golgi
3. Explain the use as a marker

1. Exported Proteins
2. Every known cell has the same function for RER
3. Excretory cells are polarized
   1. most organelles smooshed to basal end
   2. vesicles at apical end

1. Very tubular/Not flat
2. remove hydrophobic "invaders"

   Annotations:

   • This process can be problematic though. It can sometimes form molecules that are just as or more dangerous than the hydrophobic predecessors. Ex. Large molecules, once oxygenated, may become very flat. here flattened molecules may slide between base pairs in DNA and will be carcinogenic. Why hasn't this faulty trait been weened out of the population through natural selection? The answer is simple, traits are only selected during the pre-reproductive phase. Cancer is a disease that appears later in life. Therefore, it will not affect pre-reproductive populations
   1. Detox
3. Varying functions
   1. Ca2+ ions
   2. Synth. Steroid hormones, cholesterol
   3. synth. of hormones

Annotations:

• Proteins to be secreted contain a specialized amino acid sequence in the first few amino acids. This is called the signal sequence.