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lecture 22&23

lecture 22&23 - LECTURES 22 23 16 21 Oct 2009(P J...

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LECTURES 22 & 23 16 & 21 Oct 2009 (P. J. Hollenbeck) BIOL 231 Protein Synthesis on the Endoplasmic Reticulum Read: Chap. 15: 495-509; Figs. 15-12 to 15-17; DVD 15.3, 15.4, 15.7, 15.13, 15.15 Problems: 62-68 (not due until 10/28) + Exam III’04 #1; Exam III'07 #3 <We return to translation so we can understand how proteins get into all these membranes and organelles that we’ve seen. In particular, how do proteins end up in the right compartment?> I. Translation in the cell A. Emerging from the ribosome (recall the march of ribosomes along the mRNA) (1) The N-terminus of the protein emerges from the ribosome and can begin folding even as the rest of the protein is still being synthesized. Thus, although we’ve said that the 3-dimensional structure of a protein results directly from the order of its amino acids (primary structure), they do not all have opportunity to interact simultaneously. (2) Local folding (secondary structure) is therefore likely to precede the final formation of the full tertiary structure of the protein. (3) Thus, denatured proteins (unfolded by conditions into a random coil) often cannot re-fold properly on their own, in part because there is no natural pathway of folding that starts with a completely synthesized, “unwound” polypeptide. B. Location of proteins (consider the boundaries) (1) A sticking point: mRNA is found in the cytosol; but proteins are not only located in the cytosol, but also within all of the membranes of the cell, as we’ve discussed. In addition, many proteins reside within the lumen of the endomembrane system, a distinct space with a boundary between it and the cytosol. And some end up leaving the cell entirely. (2) How do they get there? The “targeting” information, like the protein structure itself, resides in the amino acid sequence [Gunter Blobel, Nobel prize 1999]. But what are the mechanisms of protein traffic? C. Membrane budding & fusion (how do proteins get to the cell surface?) (1) The secretory pathway: proteins that will be secreted from the cell must get into the lumen of the endomembrane system, then be budded off into secretory vesicles. The latter must then fuse with the plasma membrane to release the proteins to the extracellular milieu. A similar same path is followed by proteins destined to reside in the plasma membrane, from the ER membrane to a vesicle membrane to the surface. (2) This is accomplished by coupling the synthesis of proteins to their transport across the membrane of the endoplasmic reticulum. <OK, how is this organized, & how does it lead to secretion? > -1-
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II. Protein synthesis on the ER A. Pulse-chase expts with intact cells (choice of cell important for clear result) [Fig at right] (1) Jamieson & Palade (Nobel laureate, 1974) performed “pulse” radiolabeling of pancreatic secretory cells and then fixed them for EM autoradiography at intervals up to an hour or so. This provided a series of snapshots of the location of proteins that had been synthesized at a narrow window in time.
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lecture 22&amp;23 - LECTURES 22 23 16 21 Oct 2009(P J...

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