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Prion biology problem Prion biology problem space:space:
Mad cows, itchy sheep and Mad cows, itchy sheep and protein structureprotein structure
Touring the problem spaceTouring the problem space
I.I. History and introduction to prion biologyHistory and introduction to prion biology
II.II. Idea of conformational changeIdea of conformational change
III.III. Exploring sequences of prion proteinsExploring sequences of prion proteins
IV.IV. Exploring physical properties and Exploring physical properties and structures of prion proteinsstructures of prion proteins
V.V. Linking evolution, structure and function Linking evolution, structure and function of prion proteinsof prion proteins
Touring the problem spaceTouring the problem space
I.I. History and introduction to History and introduction to prion biologyprion biology
II.II. Idea of conformational change Idea of conformational change III.III. Exploring sequences of prion proteinsExploring sequences of prion proteinsIV.IV. Exploring physical properties and Exploring physical properties and
structures of prion proteinsstructures of prion proteinsV.V. Linking evolution, structure and function Linking evolution, structure and function
of prion proteinsof prion proteins
Prion HistoryPrion History
1717thth ct. “Sheep are strangely dizzy” – scrapie described ct. “Sheep are strangely dizzy” – scrapie described in Europein Europe
1950’s: Carleton Gajdusek studied cannibalistic 1950’s: Carleton Gajdusek studied cannibalistic ceremonies of Fore tribe in New Guinea and discovered ceremonies of Fore tribe in New Guinea and discovered the Kuru diseasethe Kuru disease
Kuru attacks dura matter of brain and creates vacuoles, Kuru attacks dura matter of brain and creates vacuoles, causing infected person to deteriorate and die within 3 causing infected person to deteriorate and die within 3 monthsmonths
Prion History
1960: Alper - DNA destroying UV radiation doesn’t prevent scrapie infections.
1966: Gajdusek - Chimps get Kuru from ingesting infected brain tissue.
1967: Griffith - Speculated that TSE’s due to protein conformational changes.
1981: Mertz - Fibrils detected in scrapie infected mouse brains. (CONTINUED…)http://www.css.edu/USERS/pstein/CHM3430/11
1700s: Scrapie behavior described
1997: Stanley Prusiner proposed a link between kuru, CJD, 1997: Stanley Prusiner proposed a link between kuru, CJD, scrapie and BSE. scrapie and BSE.
-Proposed that “prion” was responsible for spongiform -Proposed that “prion” was responsible for spongiform manifestations in BSE-infected patientsmanifestations in BSE-infected patients
DEFINITION: DEFINITION:
PRION = Proteinaceous Infectious ParticlesPRION = Proteinaceous Infectious Particles
Prion protein exists in two forms:Prion protein exists in two forms:
PrPc vs. PrPscPrPc vs. PrPsc
http://www.mentorcorp.com/gynecology/prion_history.htmhttp://www.mentorcorp.com/gynecology/prion_history.htmhttp://www.css.edu/USERS/pstein/CHM3430/Prions.ppthttp://www.css.edu/USERS/pstein/CHM3430/Prions.ppt
Protein folding as we know itProtein folding as we know it
http://images.google.com/imgres?imgurl=press2.nci.nih.gov/sciencebehind/snps_cancer/snps_cancer/images/21.jpg&imgrefurl=http://press2.nci.nih.gov/sciencebehind/snps_cancer/snps_cancer/snps_cancer21.htm&h=461&w=402&sz=22&tbnid=32gDooND7ssJ:&tbnh=124&tbnw=109&start=1&prev=/images%3Fq%3Dprotein%2Bfolding%26hl%3Den%26lr%3D%26ie%3DUTF-8%26sa%3DG
Touring the problem spaceTouring the problem space
I.I. History and introduction to prion biologyHistory and introduction to prion biology
II.II. Idea of conformational changeIdea of conformational changeIII.III. Exploring sequences of prion proteinsExploring sequences of prion proteins
IV.IV. Exploring physical properties and Exploring physical properties and structures of prion proteinsstructures of prion proteins
V.V. Linking evolution, structure and function Linking evolution, structure and function of prion proteinsof prion proteins
Prion protein foldingPrion protein folding
PrPcPrPsc
kfolding-c
kfolding-sc
kunfolding-sckunfolding-c
Intermediates?
Prion folding model as link between Prion folding model as link between chemistry and biologychemistry and biology
What do folding rates and equilibrium constants What do folding rates and equilibrium constants mean?mean?
If a protein has stability of 7 kcal mol-1, how many molecules are unfolded at 37oC?
Would you expect PrP to be more or less stable than other proteins?
Folded UnfoldedKeq
One protein: Two structuresOne protein: Two structures
http://www.uccs.edu/~rmelamed/MicroFall2002/Chapter%2010/Prion%20Structure.htmlhttp://www.uccs.edu/~rmelamed/MicroFall2002/Chapter%2010/Prion%20Structure.html
PrPc“NORMAL”
conformation
PrPsc“BAD”
conformation
Not all pretty pictures can be Not all pretty pictures can be trusted!trusted!
Based on NMR results Proposed
based on low-resolution structural studies
Disease transmission modesDisease transmission modes
Infections Infections (Kuru, scrapie, bovine spongiform (Kuru, scrapie, bovine spongiform encephalophathies, etc)encephalophathies, etc)
InheritedInherited (Creutzfeld-Jacob disease, (Creutzfeld-Jacob disease, Gerstmann-Straussler-Scheinker disease, Fatal Gerstmann-Straussler-Scheinker disease, Fatal familial insomnia)familial insomnia)
SporadicSporadic (BSE, CJD ?) (BSE, CJD ?)
Link to Link to The Case of the Cherry Hill ClusterThe Case of the Cherry Hill Cluster By D.T. MAXBy D.T. MAX
NY Times magazinePublished: March 28, 2004 NY Times magazinePublished: March 28, 2004
Prion diseases and Koch’s Prion diseases and Koch’s postulatespostulates
I.I. The agent must be present in every case of the The agent must be present in every case of the diseasedisease
II.II. The agent must be isolated from the host and grown The agent must be isolated from the host and grown in a lab dishin a lab dish
III.III. The disease must be reproduced when a pure The disease must be reproduced when a pure culture of the agent is inoculated into a healthy culture of the agent is inoculated into a healthy susceptible hostsusceptible host
IV.IV. The same agent must be recovered again from the The same agent must be recovered again from the
experimentally infected hostexperimentally infected host..
Can this be applied to understanding prion diseases?
Touring the problem spaceTouring the problem space
I.I. History and introduction to prion biologyHistory and introduction to prion biologyII.II. Idea of conformational changeIdea of conformational change
III.III. Exploring sequences of prion Exploring sequences of prion proteinsproteins
IV.IV. Exploring physical properties and Exploring physical properties and structures of prion proteinsstructures of prion proteins
V.V. Linking evolution, structure and function Linking evolution, structure and function of prion proteinsof prion proteins
Multiple ways to visualize and Multiple ways to visualize and analyze protein sequenceanalyze protein sequence
Use GeneDoc from Pittsburgh Use GeneDoc from Pittsburgh Supercomputing Center to analyze Supercomputing Center to analyze multiple prion sequencesmultiple prion sequences
By using different coloring schemes, can By using different coloring schemes, can visualize and visualize and quantifyquantify conservation of conservation of different propertiesdifferent properties
Comparison of human, cow, sheep Comparison of human, cow, sheep and mouse prion protein sequenceand mouse prion protein sequence
Conservarion of polar vs. non-polar Conservarion of polar vs. non-polar patternpattern
PROPERTIESLevel 1 DEHKRNQT POLARLIVMFYWAGCP NON-POLAR
Level 2DEHKRNQSTLIVMFYWAG
Analyzing charge distributionAnalyzing charge distribution
Level 2DEHKR CHARGEDNQST POLAR UNCHARGEDLIVMFYW HYDROPHOBICAG SMALL hydrophobic
Examples of questions that can be Examples of questions that can be examined through sequence examined through sequence
analysesanalysesDetermining evolutionary relationships Determining evolutionary relationships
among the various organisms examined.among the various organisms examined. Investigating how amino acid sequence Investigating how amino acid sequence
may be linked to the overall structure of may be linked to the overall structure of the proteinthe protein
Examining the role of repetitive elements Examining the role of repetitive elements in prion homologies. in prion homologies.
Touring the problem spaceTouring the problem space
I.I. History and introduction to prion biologyHistory and introduction to prion biologyII.II. Idea of conformational changeIdea of conformational changeIII.III. Exploring sequences of prion proteinsExploring sequences of prion proteins
IV.IV. Exploring physical properties Exploring physical properties and structures of prion and structures of prion proteinsproteins
V.V. Linking evolution, structure and function Linking evolution, structure and function of prion proteinsof prion proteins
Exploring physical characteristicsExploring physical characteristics
How big is the prion protein? How big is the prion protein?
How does that compare to other proteins?How does that compare to other proteins?
Where is it localized in the cell?Where is it localized in the cell?
Is its size or shape unusual?Is its size or shape unusual?
Is the distribution of amino acids unusual?Is the distribution of amino acids unusual?
PROTPARAM: a tool for analysis of PROTPARAM: a tool for analysis of physical propertiesphysical properties
http://ca.expasy.org/cgi-bin/protparamhttp://ca.expasy.org/cgi-bin/protparamNumber of amino acids:Number of amino acids: 253 253 Molecular weight:Molecular weight: 27629.1 27629.1
Theoretical pI:Theoretical pI: 9.23 9.23
Amino acid composition:Amino acid composition: Ala (A) 10 4.0% Arg (R) 11 Ala (A) 10 4.0% Arg (R) 11 4.3% Asn (N) 12 4.7% Asp (D) 6 2.4% Cys (C) 4 1.6% 4.3% Asn (N) 12 4.7% Asp (D) 6 2.4% Cys (C) 4 1.6% Gln (Q) 14 5.5% Glu (E) 9 3.6% Gly (G) 45 17.8% His Gln (Q) 14 5.5% Glu (E) 9 3.6% Gly (G) 45 17.8% His (H) 10 4.0% Ile (I) 9 3.6% Leu (L) 12 4.7% Lys (K) 11 (H) 10 4.0% Ile (I) 9 3.6% Leu (L) 12 4.7% Lys (K) 11 4.3% Met (M) 11 4.3% Phe (F) 7 2.8% Pro (P) 17 6.7% 4.3% Met (M) 11 4.3% Phe (F) 7 2.8% Pro (P) 17 6.7% Ser (S) 15 5.9% Thr (T) 13 5.1% Trp (W) 9 3.6% Tyr (Y) Ser (S) 15 5.9% Thr (T) 13 5.1% Trp (W) 9 3.6% Tyr (Y) 13 5.1% Val (V) 15 5.9% 13 5.1% Val (V) 15 5.9%
Secondary structural elements can Secondary structural elements can be analyzed in isolationbe analyzed in isolation
Helices can be dissected by Helices can be dissected by mapping onto helical wheelmapping onto helical wheel
http://bioinf.man.ac.uk/%7Egibson/HelixDraw/helixdraw.html
Examples of questions that can be Examples of questions that can be explored through structural explored through structural
analysesanalysesSome mutations have been associated with Some mutations have been associated with
prion diseases. By mapping these onto prion diseases. By mapping these onto structures, propose a hypothesis to structures, propose a hypothesis to explain this.explain this.
Most of the variation seems to be in the N-Most of the variation seems to be in the N-terminal region. Is there homology to this terminal region. Is there homology to this region in other proteins?region in other proteins?
Touring the problem spaceTouring the problem space
I.I. History and introduction to prion biologyHistory and introduction to prion biology
II.II. Idea of conformational changeIdea of conformational change
III.III. Exploring sequences of prion proteinsExploring sequences of prion proteins
IV.IV. Exploring physical properties and Exploring physical properties and structures of prion proteinsstructures of prion proteins
V.V. Linking evolution, structure Linking evolution, structure and function of prion proteinsand function of prion proteins
What does PrP do?
•GPI- anchored cell surface protein
•Knockout mice mostly normal
•Might be involved in some signaling cascades
•Interaction with hypothetical “protein x” might be needed for structural conversion
What makes prion a prion?What makes prion a prion?
Why don’t other proteins adopt two Why don’t other proteins adopt two drastically different stably folded drastically different stably folded conformations?conformations?
A helix is a helix…but why are helices in A helix is a helix…but why are helices in prion protein likely to undergo a big prion protein likely to undergo a big structural change?structural change?
How would you test/further explore these questions?
““Big picture”Big picture” questions as a questions as a platform for development of platform for development of
testable hypothesestestable hypotheses
Why would nature evolve prions?Why would nature evolve prions?
How would you tell a prion if you saw one?How would you tell a prion if you saw one?
Yeast prions do not cause a disease. Are Yeast prions do not cause a disease. Are there other non-harmful prion-like there other non-harmful prion-like proteins? Can prions be beneficial?proteins? Can prions be beneficial?