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Protein Trafficking Figure 5 Paper

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Alexis Evans 10.16.2019 The Synthesis and Posttranslational Fate of a Bee Venom Protein Source: Ibrahimi, I. 1987. Signal recognition particle causes a transient arrest in the biosynthesis of prep ... romelittin and mediates its translocation across mammalian endoplasmic reticulum. J. Cell Biol. 104: 61–66. Corresponding chapter(s) in the textbook: Chapter 11 Review the following terms before working on the problem: secretion, protein synthesis, posttranslational protein processing, in vitro translation, mRNA, [3H]proline/ [35S]methionine labeling, microsomal fraction, endoplasmic reticulum, signal recognition particle (SRP), SRP receptor, proteases, SDS-polyacrylamide gel electrophoresis, autoradiography. Read the paper and answer the questions below that refers to the data described in Figure 5 of the paper. Be prepared to discuss the other experiments described, in class. EXPERIMENT Promelittin (PM) is a small secretory protein, a predominant component of queen bee venom. Its synthesis and posttranslational metabolism was studied in an in vitro translation system containing total bee venom gland mRNA, wheat germ lysate, [3H]proline, [35S]methionine, and other molecules required for protein synthesis. Supplements were added to this system as follows: Sample 1: rough microsomal fraction (RM) Sample 2: no additional components Sample 3: cycloheximide (a protein synthesis inhibitor) was added at the end of translation, followed by the addition of RM fraction and another 30 minute incubation Sample 4: rough microsomes (RM-E) treated with N-ethylmaleimide (an inhibitor of SRP receptor) were added at the beginning of translation Sample 5: same as sample 4, except this sample was digested with proteinase K after translation The samples were subjected to SDS polyacrylamide gel electrophoresis followed by autoradiography [Show More]

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