Transcript Slide 1

Prion biology problem
space:
Mad cows, itchy sheep and
protein structure
Touring the problem space
I.
II.
III.
IV.
V.
History and introduction to prion biology
Idea of conformational change
Exploring sequences of prion proteins
Exploring physical properties and
structures of prion proteins
Linking evolution, structure and function
of prion proteins
Touring the problem space
I.
History and introduction to
prion biology
II.
Idea of conformational change
Exploring sequences of prion proteins
Exploring physical properties and
structures of prion proteins
Linking evolution, structure and function
of prion proteins
III.
IV.
V.
Prion History

17th ct. “Sheep are strangely dizzy” – scrapie described
in Europe

1950’s: Carleton Gajdusek studied cannibalistic
ceremonies of Fore tribe in New Guinea and discovered
the Kuru disease

Kuru attacks dura matter of brain and creates vacuoles,
causing infected person to deteriorate and die within 3
months
Prion History
1700s: Scrapie behavior described
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

1997: Stanley Prusiner proposed a link between
kuru, CJD, scrapie and BSE.
-Proposed that “prion” was responsible for
spongiform manifestations in BSE-infected
patients
DEFINITION:
PRION = Proteinaceous Infectious Particles
Prion protein exists in two forms:
 PrPc
vs. PrPsc
Protein folding as we know it
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Touring the problem space
I.
History and introduction to prion biology
II.
Idea of conformational change
III.
Exploring sequences of prion proteins
Exploring physical properties and
structures of prion proteins
Linking evolution, structure and function
of prion proteins
IV.
V.
Prion protein folding
kfolding-c
kfolding-sc
kunfolding-c
kunfolding-sc
PrPc
Intermediates?
PrPsc
Prion folding model as link between
chemistry and biology
What do folding rates and equilibrium constants
mean?
Folded
Keq
Unfolded
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?
One protein: Two structures
PrPc
PrPsc
“NORMAL”
“BAD”
conformation
conformation

http://www.uccs.edu/~rmelamed/MicroFall2002/Chapter%2010/Prion%20Structure.html
Not all pretty pictures can be
trusted!
Based
on NMR
results
Proposed
based on
lowresolution
structural
studies
PrPc
PrPc
PrPc
PrPsc
PrPc
PrPc
PrPc
PrPc
PrPc
PrPc
PrPc
PrPc
PrPc
PrPC
PrPSC
PrPC
PrPsc fibrils
Plaque
Disease transmission modes

Infections (Kuru, scrapie, bovine spongiform
encephalophathies, etc)

Inherited (Creutzfeld-Jacob disease,
Gerstmann-Straussler-Scheinker disease, Fatal
familial insomnia)

Sporadic (BSE, CJD ?)


Link to The Case of the Cherry Hill Cluster
By D.T. MAX
NY Times magazinePublished: March 28, 2004
Prion diseases and Koch’s
postulates
I.
II.
III.
IV.
The agent must be present in every case of the
disease
The agent must be isolated from the host and grown
in a lab dish
The disease must be reproduced when a pure
culture of the agent is inoculated into a healthy
susceptible host
The same agent must be recovered again from the
experimentally infected host.
Can this be applied to
understanding prion diseases?
Touring the problem space
I.
II.
History and introduction to prion biology
Idea of conformational change
III.
Exploring sequences of prion
proteins
IV.
Exploring physical properties and
structures of prion proteins
Linking evolution, structure and function
of prion proteins
V.
Global view of the amino acid
sequence of PrP
Multiple ways to visualize and
analyze protein sequence
Use GeneDoc from Pittsburgh
Supercomputing Center to analyze
multiple prion sequences
 By using different coloring schemes, can
visualize and quantify conservation of
different properties

Comparison of human, cow, sheep
and mouse prion protein sequence
Conservarion of polar vs. non-polar
pattern
PROPERTIES
Level 1
DEHKRNQT
LIVMFYWAGCP
POLAR
NON-POLAR
Analyzing charge distribution
Level 2
DEHKR
NQST
LIVMFYW
AG
CHARGED
POLAR UNCHARGED
HYDROPHOBIC
SMALL hydrophobic
Expanded amino acid properties
Examples of questions that can be
examined through sequence
analyses
Determining evolutionary relationships
among the various organisms examined.
 Investigating how amino acid sequence
may be linked to the overall structure of
the protein
 Examining the role of repetitive elements
in prion homologies.

Touring the problem space
I.
II.
III.
History and introduction to prion biology
Idea of conformational change
Exploring sequences of prion proteins
IV.
Exploring physical properties
and structures of prion
proteins
V.
Linking evolution, structure and function
of prion proteins
Exploring physical characteristics
How big is the prion protein?
How does that compare to other proteins?
Where is it localized in the cell?
Is its size or shape unusual?
Is the distribution of amino acids unusual?
PROTPARAM: a tool for analysis of
physical properties

http://ca.expasy.org/cgi-bin/protparam
Number of amino acids: 253
Molecular weight: 27629.1
Theoretical pI: 9.23
Amino acid composition: 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%
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
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)
13 5.1% Val (V) 15 5.9%
Visualizing protein structure with
Cn3D
Secondary structural elements can
be analyzed in isolation
Helices can be dissected by
mapping onto helical wheel
http://bioinf.man.ac.uk/%7Egibson/HelixDraw/helixdraw.html
Examples of questions that can be
explored through structural
analyses
Some mutations have been associated with
prion diseases. By mapping these onto
structures, propose a hypothesis to
explain this.
Most of the variation seems to be in the Nterminal region. Is there homology to this
region in other proteins?
Touring the problem space
I.
II.
III.
IV.
V.
History and introduction to prion biology
Idea of conformational change
Exploring sequences of prion proteins
Exploring physical properties and
structures of prion proteins
Linking evolution, structure
and 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?

Why don’t other proteins adopt two
drastically different stably folded
conformations?

A helix is a helix…but why are helices in
prion protein likely to undergo a big
structural change?
How would you test/further
explore these questions?
“Big picture” questions as a
platform for development of
testable hypotheses
Why would nature evolve prions?
How would you tell a prion if you saw one?
Yeast prions do not cause a disease. Are
there other non-harmful prion-like
proteins? Can prions be beneficial?