Natural Sciences > Biology > Protein Folding, Misfolding and Human Disease

Protein Folding, Misfolding and Human Disease posted by duggu on 12/8/2007

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Abstract/Syllabus

Courseware/Lectures

Test/Tutorials

Course Highlights

This course features a full bibliography in the readings section.

Course Description

This course is one of many Advanced Undergraduate Seminars offered by the Biology Department at MIT. These seminars are tailored for students with an interest in using primary research literature to discuss and learn about current biological research in a highly interactive setting. The instructor for this course, Dr. Kosinski-Collins, is a member of the HHMI Education Group.

Maintenance of the complex three-dimensional structure adopted by a protein in the cell is vital for function. Oftentimes, as a consequence of environmental stress, genetic mutation, and/or infection, the folded structure of a protein gets altered and multiple proteins stick and fall out of solution in a process known as aggregation. In many protein aggregation diseases, incorrectly folded proteins self-associate, forming fiber-like aggregates that cause brain cell death and dementia. In this course, the molecular and biochemical basis of the prion diseases, which include bovine spongiform encephalopathy (mad cow disease), Creutzfedt-Jakob disease and kuru will be examined. Also discussed are other classes of misfolding diseases such as Alzheimer's disease and Huntington's disease. The proteins involved in all of these disorders and how the proteins' three dimensional structures change during the course of these afflictions is covered as well as why prions from certain species cannot infect animals from other species based on protein sequence and structure. The course will then address possible detection methods and therapies that are under development to treat some of the protein aggregation diseases.

Syllabus

Course Format

During the typical class session, we will analyze two papers in a discussion-based format. Everyone should read both papers before class, and each student should be prepared to answer questions. At the beginning of each class, we will have a question and answer period in which the instructor will respond to any specific questions that the students may have thought of while reading the articles. Immediately following, one or two students will present a brief summary of techniques essential to the topic of the papers. Students may volunteer to present the technique(s) of interest during the week prior to the discussion. We will spend the remainder of the class discussing the data presented in the article by analyzing each figure and resulting conclusions. Each class will conclude with a brief outline of the following week's class introduced by the instructor.

Note: The review and response articles have been included as supplemental material. I encourage you to read them to get a feel for the topic background, but they are not required reading and we will not discuss them directly.

Expectations

Because this course has no graded exams, students are expected to attend every class meeting. The major focus will be on discussion and interpretation of scientific papers, so student attendance and participation is essential. Other than week 1, there will be no formal lectures and students should come to class prepared to participate in a discussion of the assigned papers. Missing a class should occur only in extreme circumstances. If a student knows he/she must miss a class, he/she must contact the instructor in advance and receive permission for the absence. Students who have been allowed an excused absence will be asked to complete a written assignment concerning the papers discussed during the missed class session.

Prerequisites

As the goal of this course is to familiarize students with the reading and critical evaluation of the primary literature in the protein folding field, students are expected to have completed 7.03 (Genetics), 7.05 (Biochemistry), 7.06 (Cell Biology) or 7.08 (Molecular Biology). We will be dealing with several topics related to protein structure and design, and so a reasonable knowledge of introductory chemistry or biochemistry will be helpful.

Grading

There are no formal exams. The class has a pass/fail grading system.

Assignments

In addition to the weekly reading assignments, there will be three one to two page written assignments and an oral presentation coinciding with the first written assignment.

Calendar

WEEK #	TOPICS	KEY DATES
1	Background of Protein Misfolding and Disease
2	Origins of the Protein-only Hypothesis of Prions
3	Controversial Evidence against the Virus-hypothesis of Prion
4	Pathology of the Transmissible Spongiform Encephalopathies
5	Structure of the Prion Protein
6	The Prion Species Barrier	Assignment 2 due
7	Detection of Prion Diseases in Humans and Animals
8	Alzheimer's Disease
9	Transmission Electron Microscopy: Field trip to TEM facility
10	Oligomeric Ring Structures in Alzheimer's and Parkinson's Disease
11	Parkinson's Disease and α-synuclein
12	Polyglutamine repeats in Kennedy's and Huntington's Diseases
13	Senile Systematic Amyloidosis and Inflammation	Assignment 3 proposal due

Readings

The Techniques listed in the table below are topical examples of the first writing assignment. Some downloadable handouts are available and describe in greater detail the experimental methods and technologies used in the assigned literature. These handouts are included courtesy of the students listed in the Readings column.

WEEK #	TOPICS	OVERVIEWS	READINGS
1	Background of Protein Misfolding and Disease	During the first class session, the instructor and the students will introduce themselves. The instructor will introduce the course and go over requirements and expectations outlined in the syllabus. The instructor then will present a brief lecture outlining the background and basics of protein folding, aggregation, and disease. Finally, the topic of week 2 will be introduced and the technique for week 2 will be assigned to one student.	Review Articles Kelly, J. W. "Alternative Conformations of Amyloidogenic Proteins Govern their Behavior." Curr. Opin. Struct. Biol. 6 (1996): 11-17. Ross, C. A., and M. A. Poirer. "Protein Aggregation and Neurodegenerative Disease." Nature Med. 10 (2004): s10-s17.
2	Origins of the Protein-only Hypothesis of Prions	Scrapie was originally observed and documented by farmers in their sheep flocks in the early 1900's. Shepards would notice a strange behavior among a few members of their flock that ultimately resulted in dementia and death. Scientists identified a transmissible element in brain from affected sheep soon afterward, although the exact composition from this "infectious" particle would remain unproven for years afterward.	Pattison, I. H., and K. M. Jones. "Detection of the Scrapie Agent in Tissues of Normal Mice and in Tumours of Tumour-bearing but Otherwise Normal Mice." Nature 217 (1968): 102-104. Response to (1): Mackay, J. M. "Detection of Scrapie Agent in Tissue of Normal Mice." Nature 219 (1968): 182-183. Clark, M. C., and D. A. Haig. "Evidence for the Multiplication of Scrapie Agent in Cell Culture." Nature 225 (1970): 100-101. Techniques Protein Purification and Column Chromatography: Size Exclusion, Cation/Anion/Hydrophobic Exchange.
3	Controversial Evidence against the Virus-hypothesis of Prion	This week we will discuss Stanley Prusiner's early work on scrapie for which he would eventually win the Nobel Prize. Although the concept of a transmissible protein element still remained unproven definitively with these articles, Prusiner and his colleagues had finally shown that material from scrapie infected mice could infect other animals.	Review: Prusiner, S. B. "Novel Proteinaceous Infectious Particles Cause Scrapie." Science 216 (1982): 136-144. Prusiner, S. B., M. P. McKinley, D. F. Groth, K. A. Bowman, N. I. Mock, S. P. Cohran, and F. Masiarz. "Scrapie Agent Contains a Hydrophobic Protein." Proc. Natl. Acad. Sci. 78 (1981): 6675-6679. Diener, T. O., M. P. McKinley, and S. B. Prusiner. "Viroids and Prions." Proc. Natl. Acad. Sci. USA 79 (1982): 5220-5224. The Final Proof Legname, G., I. V. Baskakov, H. O. Nguyen, D. Rienser, F. E. Cohen, S. J. DeArmond, and S. B. Prusiner. "Synthetic Mammalian Prions." Science 305 (2004): 673-676. Science Magazine Techniques Polyacrylamide Gel Electrophoresis (SDS and Native). Purification: Density Gradient Centrifugation (Sucrose & CsCl).
4	Pathology of the Transmissible Spongiform Encephalopathies	After Prusiner's initial publications, Creutzfeldt-Jakob disease and kuru were soon classified as a prion diseases. Scientists began examining the brains of patients who had died from these illnesses to see if they could identify any common pathologies. This week, we will look at the plaques found in the brains of prion-infected animals.	Tranchant, C., L. Geranton, C. Guirand-Chaumeil, M. Mohr, and J. M. Warter. "Basis of Phenotypic Variability in Sporadic Creutzfeldt-Jakob Disease." Neurology 52 (1999): 1244-1249. Karmysheva, V. Y., V. V. Pogodina, and V. M. Roikhel. "Cytopathological Changes in Human and Animals Brains in Prion Diseases." Neurosci Behav Physiol. 34 (2004): 509-513. Neuroscience and Behavioral Physiology Techniques Transmission Electron Microscopy/Scanning Electron Microscopy (PDF) (Courtesy of anonymous student. Used with permission.)
5	Structure of the Prion Protein	Prion diseases were eventually classified as protein misfolding disorders. However, a fundamental problem still remained, how are the proteins misfolded in the scrapie conformation? We will look at two studies that address the 3-dimensional conformation of the native prion protein and the infectious scrapie state using cryoEM and X-ray crystallography.	Goverts, C., H. Wille, S. B. Prusiner, and F. E. Cohen. "Evidence for the Assembly of Prions with Left-handed Beta Helices into Trimers." Proc. Natl. Acad. Sci. 101 (2004): 8342-8347. Wille, H., M. D. Michelitsch, V. Guenebaut, S. Supattapone, A. Serban, F. E. Cohen, D. A. Agard, and S. B. Prusiner. "Structural Studies of the Scrapie Prion Protein by Electron Crystallography." Proc. Natl. Acad. Sci. USA 99 (2002): 3563-3568. Techniques X-ray Crystallography. Nuclear Magnetic Resonance (PDF) (Courtesy of John Cassady. Used with permission.)
6	The Prion Species Barrier	Possible one of the most remarkable mysterious of prion diseases is the relative difficult involved in transferring the scrapie form from one species to another. We will discuss how, where, and why this species barrier exists between even closely related animals. Discussion with Dr. Peter Chien.	Raymond, G., A. Bossers, L. Raymond, K. O'Rourke, L. McHolland, P. Bryant, M. Miller, E. Williams, M. Smits, and B. Caughey. "Evidence of a Molecular Barrier Limiting Susceptibility of Humans, Cattle and Sheep to Chronic Wasting Disease." EMBO J. 19 (2000): 4425-4430. The EMBO Journal Pertez, D., M. Scoot, D. Groth, R. Williamson, D. Burton, F. E. Cohen, and S. B. Prusiner. "Strain-specific Relative Conformational Stability of the Scrapie Prion Protein." Protein Sci. 10 (2001): 854-863.
7	Detection of Prion Diseases in Humans and Animals	One major problem in the battle against the prion diseases is that it has been exceedingly difficult to develop a reproducible, accurate detection assay for infected animals. Although a human or animal may contract a prion disease early in life, the symptoms of dementia may not be revealed until years afterward. For this reason, an infected herd of cows may be eaten by humans or used as feed long before the BSE is ever even detected. We will discuss two novel approaches to detecting the presence of the diseases.	Saborio, G. P., B. Permanne, and C. Soto. "Senstitve Detection of Pathological Prion Protein by Cyclic Ampliflication of Protein Misfolding." Nature 411 (2001): 810-813. Nature publishing group Thackray, A. M., J. Y. Madec, E. Wong, R. Morgan-Warren, D. R. Brown, T. Baron, and R. Bujdoso. "Detection of BSE, Ovine Scrapie Prion-related Protein (PrPSc) and Normal PrPc by Monoclonal Antibodies Raised to Copper Refolded Proteins." Biochem J. 370 (2003): 81-90. Techniques Monoclonal and Polyclonal Antibodies (PDF) (Courtesy of Alex Bagley. Used with permission.)
8	Alzheimer's Disease	Alzheimer's disease is characterized by the formation of long amyloid filaments in the brains of infected patients. These filaments are composed of varying amounts of the Aß-peptide. We will examine how the Aß-peptide is released into the neurons of Alzheimer's infected mice. We will also look at the mechanism of fiber formation of Aß identified in vitro using atomic force microscopy.	Harper, J. D., C. M. Lieber, and P. T. Lansbury, Jr. "Atomic Force Microscopic Imaging of Seeded Fibril Formation and Fibril Branching by the Alzheimer's Disease Amyloid-beta Protein." Chem. Biol. 4 (1997): 951-959. Borchelt, D. R., T. Ratovitski, J. Van Lare, M. K. Lee, V. Gonzales, N. A. Jenkins, N. G. Copeland, D. L. Price, and S. S. Sisodia. "Accelerated Amyloid Deposition in the Brains of Transgenic Mice Coexpressing Mutant Presenilin 1 and Amyloid Precursor Proteins." Neuron 19 (1997): 939-945. Techniques Atomic Force Microscopy.
9	Transmission Electron Microscopy	Introduction by Dr. Peter Weigele
10	Oligomeric Ring Structures in Alzheimer's and Parkinson's Disease	A relatively novel theory about Alzheimer's Aß-peptide toxicity has been introduced to the scientific community. We now think that it may not be the amyloid fiber filaments that cause neuron death and disease onset. Small oligomeric ring structures may disrupt ion concentrations in brain cells and induce cytotoxicity. This week, we will address ion fluctuations occurring during amyloidogenic neuron death and the observed structures of cytotoxic pores species.	Kawahara, M., and Y. Kuroda. "Molecular Mechanism of Neurodegeneration Induced by Alzheimer's β-amyloid Protein: Channel Formation and Disruption of Calcium Homeostasis." Brain Res. Bul. 53 (2000): 389-397. Lashuel, H. A., B. M. Petre, J. Wall, M. Simon, R. J. Nowak, T. Waltz, and P. T. Lansbury, Jr. "Alpha-synuclein, especially the Parkinson's Disease Associated Mutants, Form Pore-like Annular and Tubular Protofibrils." J Mol Biol. 322 (2002): 1089-1102.
11	Parkinson's Disease and α-synuclein	This week we will change our focus from Alzheimer's disease to α-synuclein and Parkinson's. We will examine structures of the infectious α-synuclein particles and possible mechanisms of aggregate formation and toxicity.	Park, J. Y., P. T. Lansbury, Jr. "Beta-synuclein Inhibits Formation of Alpha-synuclein Protofibrils: A Possible Therapeutic Strategy against Parkinson's Disease." Biochemistry 42 (2003): 3696-3700. ACS Publications Zhu, M., J. Li, and A. L. Fink. "The Association of Alpha-synuclein with Membranes Affects Bilayer Structure, Stability, and Fibril Formation." J Biol Chem. 278 (2003): 40186-40197.
12	Polyglutamine repeats in Kennedy's and Huntington's Diseases	This week, we will turn our attention to polyglutamine expansion diseases. Successive repeats of glutamine in certain proteins will cause self-association and aggregation of the molecule. We will look at the mechanism and affect of this process in Huntington's and Kennedy's disease.	Buchanan, G., M. Yang, A. Cheong, J. M. Harris, R. A. Irvine, P. F. Lambert, N. L. Moore, M. Raynor, P. J. Neufing, G. A. Coetzee, and W. D. Tilley. "Structural and Functional Consequences of Glutamine Tract Variation in the Androgen Receptor." Hum Mol Genet. 13 (2004): 1677-92. Human Molecular Genetics Busch, A., S. Engemann, R. Lurz, H. Okazawa, H. Lehrach, and E. E. Wanker. "Mutant Huntingtin Promotes the Fibrillogenesis of Wild-type Huntingtin: A Potential Mechanism for Loss of Huntingtin Function in Huntington's Disease." J Biol Chem. 278 (2003): 41452-61.
13	Senile Systematic Amyloidosis and Inflammation	This week we will discuss two very different papers. First, we will examine the mechanism of transthyretin aggregation amyloid formation. This was one of the first proteins in which the amyloidogenic intermediate was identified and characterized in vitro . Then, we will switch modes slightly and look at how computer modeling can be used to predict protein structure and aggregation.	Colon, W., and J. W. Kelly. "Partial Denaturation of Transthyretin is Sufficient for Amyloid Fibril Formation in vitro." Biochemistry 31 (1992): 8654-8660. ACS Publications Armen, R. S., M. L. DeMarco, D. O. Alonso, and V. Daggett. "Pauling and Corey's α-pleated Sheet Structure May Define the Prefibrillar Amyloidogenic Intermediate in Amyloid Disease." Proc Natl Acad Sci. USA (2004). Techniques Fluorescence Spectroscopy. Circular Dichroism.