Lecture 7

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Lecture 7
2013-02-04 01:57:51
BIS 104

ER modification, secretory system, Golgi apparatus, vesicular tubular cluster,
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  1. ER Lumen modifications
    • a) Removal of signal sequence
    • b) Oxidation of sulfhydryl groups
    • c) N-Glycosylation
    • d) Attachment of GlycosylPhosphatidyl-Inositol (GPI) anchor (some proteins)
    • e) Protein folding by chaperones
  2. N-Glycosylation Structure

    • Squares = N-acetylglucosamine (G IN Ac)
    • Green Circles = Mannose
    • Blue Circles = Glucose
  3. N-Glycosylation coding

    Aspargine - Any - Serine or Threonine
  4. BiP - Binding Immunoglobulin Protein Role
    Recognizes incorrectly folded protein by binding to exposed AA that would normally be buried in the interior of correct polypeptide or polypeptide that have not yet been assembled into their correct oligomeric structure.
  5. Carbohydrate-binding proteins

    Example compounds?
    Class - Lectins: Bind specifically to particular carbohydrate residues

    • Calnexin
    • Calreticulin

    Binds to oligosaccharides on proteins that have not yet folded completely and keeps them in the ER until correct folding or sends them to the cytosol for degradation.
  6. Example folding procedure involving calnexin
    • 1) Unfolded protein w/ 3 glucose that undergoes glucose trimming leaving 1 glucose.
    • 2) Protein binds to Calnexin and glucosidase removes protein from Calnexin and removes glucose.
    • 3) If folded correctly, protein exits ER. If not, it binds to glucosyl transferase and gets one glucose attached to redo the cycle.
  7. Secretory System Pathway

    Because proteins utilizing secretory pathway pass ER, signals are required.

    Experiment that determined secretory system is Pulse/chase experiment by Palade.
  8. Pulse-Chase Experiment
    1. Pulse a dye or signal in a pathway for a short time.

    2. View movement of signal/dye (chase).
  9. Green fluorescent protein (GFP) fusion protein
    A method to study transit of protein through cell.

    • 1. Use genetic engineering to fuse cDNA encoding GFP to cDNA encoding protein.
    • 2. Fluorescent marker now exist on protein.
  10. Vesicular Stomatitis Virus
    • 1. Virus starts at ER
    • 2. Exits ER
    • 3. Goes through Golgi
    • 4. Moves through plasma membrane
  11. Golgi Apparatus
    Ordered series of compartments called cisternae.

  12. Transit through the secretory system is ________
    Vesicular (i.e. leaves donor compartment as budding and enters target compartment as fusion)
  13. Mature glycoproteins
    High mannose (mannose tail 1-2-2-1)

    Complex: has row of GINAc, galactose, NANA
  14. Carbohydrate units _______ from the _________ and __________ from __________
    Protrude, glycoprotein, protect, degradation
  15. Golgi apparatus layers
    • 1. Cis Golgi Network
    • 2. Cis Cisterna
    • 3. Medial Cisterna
    • 4. Trans Cisterna
    • 5. Trans Golgi Network
  16. Cis Golgi Network Role
    Sorting: Phosphorylation of oligosaccharides on lysosomal proteins.
  17. Golgi Stack Role
    Cis: Removal of MAn

    Medial: Removal of Man and addition of GlcNac

    Trans: Addition of Gal and NANA
  18. Trans Golgi Network Role
    Sorting: Sulfation of tyrosines and carbohydrates.

    Protein goes to lysosome, plasma membrane, or secretory vesicle.
  19. Vesicular tubular cluster
    Acts as cargo mover and intermediate between ER and cis Golgi network.

    COPII coated vesicle of proteins find their way to VTC.

    COPI coated vescile of proteins find their way back to ER for Golgi network
  20. Retention of resident proteins of compartments
    Soluble ER protein binds to KDEL where membrane KDEL receptors bind and keep ER proteins contained.
  21. Lysosomes function
    Main site of digestion of intracellular and extracellular debris including macromolecules and obsolete parts of the cell.
  22. Lysosome characteristics:

    Enzyme class and examples?
    Maintaining pH?
    Evolution from ______?

    • Acid Hydrolase
    • - Nucleases, proteases, lipases, glycosidase, phosphatases, sulfatases, phospholipases.

    ATP dependant hydrogen pump

    Late endosome
  23. Plant lysosome are called _______
  24. Lysosomes morphology are _________
  25. Signal Patch
    Regions of protein sequence that, when the protein is folded, signals the protein to be binded by a receptor protein in the cis-Golgi to undergo processing.

    Receptor is called GlcNAc phosphotransferase
  26. GlcNAc phosphotransferase and pathway of substrate
    Binds UDP-GlcNAc to proteins destined for lysosome and is the complex is bridged by a phophate group.

    Goes through medial and trans-Golgi where GlcNac will be removed and expose phosphorylated mannose.
  27. Golgi to Lysosome pathway
  28. Inclusion Cell Disease
    Single gene defect and recessive.

    Hydrolytic enzymes of lysosomes in fibroblast missing. Patients exhibit lots of inclusion bodies (garbage in cells).

    Elizabeth Neufeld found out that if you put defective fibroblast with regular fibroblast, defective fibroblast recovered.

    They lacked functional GlcNAc phosphotransferase so they didn't have a signal for lysosome degradation.