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1. Protein biogenesis and degradation MBB443/743 Prof. Michel R. Leroux Fall semester 2012 Wednesday & Friday 8:30 am – 10:20 am SECB 1012

2. Office hours Office South Sciences Bldg, SSB6144 Telephone 778-782-6683 e-mail leroux@sfu.ca • I will be available after the lectures in class or in my office (Wednesday and Friday) • if you have questions, please try to see me right after class • any e-mails to me should have the header “MBB443”

3. Course outline - lecture - student presentation(s) Wednesdays & Fridays - no specific tutorial time(s) assigned- no required text • total # classes scheduled: 26 • class presentations: - students give 1-2 oral presentations (in pairs if too many students) - 20 minutes each - thepaper(s) will be assigned • 1 term paper (in pairs if too many students) • 2 exams

4. Class presentations 2. introduce paper, giving enough background information for everyone to understand the paper; this should include some background information not found in the paper itself- also explain the research goals ~5 min 3. present key experimental data along with brief explanations of procedures; the data presented can be a subset of all of the paper’s data 4. explain the results obtained ~15 min Powerpoint presentations: on my computer assigned at least one week in advance Guidelines: 1. prepare a 1-page (maximum) summary of the presentation that is distributed to the class(please e-mail for distribution) If time permits: class discussion- approaches used were appropriate? - results were convincing? - further studies required?

5. Term paper based on NSERC grant application Content • topic should be related to course material • Introduction; requires literature search (~4 double-spaced pages) • research proposal and references (~6 double-spaced pages) • research proposal should describe a study that would advance our knowledge of biogenesis and/or degradation {or related topic} - describe goal, importance, procedures (brief), alternate experiments, interpretation of (anticipated) results, conclusions Format of manuscript • 10 double-spaced pages, including references • 2-2.5 cm margins all around • 12 pt Times New Roman font • references should be numbered and have the following style: Deere, J., McIntosh, A. and Crusher, W. (2000) Studies on the refolding of Ribonuclease A. Nature38, 345-368. Important ! • you must use your own words when writing the paper; in the rare case you need to use someone’s wording, use quotations and reference the paper Timeline• wait about one month before choosing your topic; report due November 16

6. Exams There are two exams • anything mentioned in class is fair game: - lectures - assigned papers (~1 for each term) - class presentations (no detailed questions) - in-class discussions • answering exam questions may also require reading a small portion of a real scientific publication • mostly short, written answers (a word, a sentence or a paragraph) • focus is on understanding, not memorization; I will tell you if there is something you should specifically memorize. • no final exam scheduled Finished on last day of class (Nov. 30) !

7. Lectures, info available on my web site www.sfu.ca/~leroux click on teaching and download lectures and other stuff (e.g. research proposal info) lowercase PASSWORD for some files: mbb-sfu

8. Grading (or 40%) (or 25%)

9. Central dogma of biology folding, assembly, targeting cellular proteins degradation (turnover) FUNCTIONAL (NATIVE) PROTEIN amino acids, peptides cellular proteins cellular proteins regulation of conformation/ function transcription translation DNA RNA PROTEIN replication

10. Link between biogenesis and degradation: ‘non-native’ proteins both processes can be grouped under the heading of quality control. Non-native (unfolded, misfolded, denatured) protein: a protein that is not properly folded and is not in a functional state • biogenesis signifies the ‘birth’ of proteins, or the transition between non-native to native states - biogenesis includes: folding, assembly, transport to and across biological membranes, refolding, chemical or structural modification • degradation represents the ‘death’ of proteins, or the transition from native to ‘non-native’ states to basic constituents - degradation includes the disposal of damaged (non-native) proteins and the timely, regulated turnover of various cellular proteins

11. Cellular processes involvingnon-native proteins: folding and assembly • proteins are synthesized on the ribosome and must fold/assemble to become native - proteins are synthesized as unfolded polypeptide chains - folding occurs co-translationally - folding (and assembly) to the native state requires the complete polypeptide chain folding assembly

12. Cellular processes involvingnon-native proteins: refolding various cellular proteins cellular stress heat/cold Native protein non-native (unfolded) protein proteotoxic chemicals aggregated protein intracellular changes

13. Cellular processes involvingnon-native proteins: transport • protein transport to, and across biological membranes - protein must be maintained in a translocation-competent state - protein must not misfold or aggregate - protein must be directed to proper membrane / cellular compartment • also: intracellular transport (to specific locations in the cytosol, nucleus, etc. (but this typically involves native proteins)

14. Cellular processes involvingnon-native proteins: regulation of protein conformation/function • under certain circumstances, the conformation (activity) of some proteins must be modulated - heat shock transcription factor is activated only during a cellular stress - steroid receptors must be kept in a form that is competent to bind ligand, but is not active - signalling molecules (kinases) are kept in an inactive conformation until phosphorylated inactive, non- native cellular event active, native

15. Cellular processes involvingnon-native proteins: degradation unfolding proteolysis proteolysis * * * amino acids peptides Protein destined for degradation antigen presentation * steps involve various cellular machineries

16. Cellular processes involvingnon-native proteins: quality control non-native protein unfolding refolding refolding degradation Native protein peptides, amino acids Native protein

17. Protein misfolding diseases

18. Protein degradation: cellular roles Degradation involves numerous cellular processes, including: processing of nascent polypeptides cleavage of proteins to peptides for antigen presentation destruction of proteins that are inefficiently folded/processed degradation of aberrant (mutant) proteins turnover of cell-cycle or other proteins that are short-lived or whose presence in the cell is strictly regulated destruction of proteins damaged due to cellular stresses (oxygen radicals, elevated/reduced temperatures, etc.) turnover of proteins that have lost activity over time - there are a large variety of proteases in the cell, and degradation is strictly controlled

19. Protein misfolding disease: amyloidosis • at least 16 different proteins are implicated in amyloid diseases • a number of different proteins can be induced to form fibrils in vitro as well amyloid formation

20. Amyloid structure electron microscopy model of filament cross-section Courtesy of Helen Saibil Dept. of Crystallography, Birkbeck College London

21. Protein modification Ubiquitination Lys degradation/other

22. Topics covered