biology_notes.docx - Cargo proteins inside donor compartment have sorting proteins that will indicate where they are supposed to go in the cell and

biology_notes.docx - Cargo proteins inside donor...

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Cargo proteins inside donor compartment have sorting proteins that will indicate where they are supposed to go in the cell and specific receptors bind terminate destination where they are supposed to go. How to study endomembrane? Pulse of radioactive amino acids. Chase in autoradiography means wait time. Different wait times when you can see where the proteins moving in different periods of time. Silver grains show up where radioactive protein has been moved to. 3 min Pulse: proteins are in the ER. Wait 20 minutes before radiographic emulsion – proteins are in the Golgi or in vesicles that are leaving Golgi. 120 minutes Chase – proteins are in the vesicles that are about to leave plasma membrane. Zymogen granules are about to leave the cell. Green fluorescent protein. Specific protein – GFP fusion of this protein by taking DNA and add the gene sequence of GFP at the end of protein. Translated protein – GFP protein is attached to carboxyl end. Subcellular fractionation. Cell-Free systems. Microsomes – functional ER and Golgi membranes that are reformed into vesicles. Further analysis, density gradient where you have all microsomes centrifuged through this density gradient. They become denser and then microsomes will be separated into smooth microsomes without ribosomes and rough microsomes with ribosomes on them. Sec mutants. Randy Shekman. No mutations manifested at the low temperature. Increase the temperature to 37 that will cause mutations in those proteins to denature and not function anymore. A – ribosome stop translation and gets broaden into trans locon and deposit proteins into lumen of the ER. Class-A mutations – cargo proteins that were supposed to be deposited into ER. B – the process of moving from ER into vesicles affected – not form vesicles, mutants accumulate in the ER or lumen of the membrane of the ER rather than moving to vesicles. C accumulate in the vesicles, D mutations in the Golgi, E stay in the secretory vesicles rather than being deposited into plasma membrane. They were able to identify the exact gene that is responsible for the defects. Endoplasmic reticulum. Rough ER and Smooth ER. Smooth ER store calcium, and it’s important for cell signaling. Proteins made by membrane-bound ribosomes are being broad into ER as they are being translated – cotranslational import. Moving into the ER as they are translated. Free ribosomes proteins they are getting imported posttranslationally. Synthesis. Gunter Blobel. Cotranslational import to make secretary proteins. SRP recognizes ER signal sequence. SRP docks ribosome onto translocon. Signal sequence interact with pore. Signal peptidase cleaves the sequence. Amino end is still inside the lumen.
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  • Fall '11
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