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Prions Gone Mad
Insight into prion diseases: structural lessons learned from fungus
Laconia SMART Team: Laura Block, Kea Schmuhl, Erin Cunningham, Tyler Foote, Ashley Toll, Cole Tidemann, Ashley Garb, Dakota Moore
Advisor: Jodie Garb, Laconia High School
Mentor: Anita Manogaran, Ph.D., Marquette University
Abstract
Recent Developments in Prion Biology
Prions in Fungi
Prion diseases, including bovine spongiform encephalopathy (mad cow) and
Creutzfeldt–Jakob disease in humans, are caused by a misfolded protein in the
brain that has the ability to convert the normal protein to the misfolded form.
Prions in the brain lead to the formation of aggregates of misfolded protein, which
are thought to be infectious and toxic to the cell. Additionally, these aggregates
are resistant to detergents or heat, and are difficult to destroy. While reliable
structural studies of human prion proteins have been unsuccessful, the structure
of the [HET-s] prion in the fungus Podospora anserina has been solved. The HET-s
protein misfolds and assembles into a higher order structure called a “solenoid,” a
tube of circular beta sheets. Laconia SMART (Students Modeling a Research Topic)
Team modeled the HET-s protein with 3D printing technology, illustrating its
solenoid structure. Each “turn” of the solenoid consists of 21 amino acids. A
pocket of hydrophobic (V244, L276, F286, W287), hydrophilic (Q240, E280), and
glycine amino acid side chains is thought to stabilize the solenoid structure. Since
human prion proteins are thought to share a similar solenoid structure with HET-s,
studying HET-s provides insight into the molecular structure of human prions,
which could potentially lead to advances in treatment of prion-related diseases.
Prion diseases are associated with protein aggregates. The aggregates form
when the normal protein undergoes misfolding (Fig. 3). This misfolded
protein has the ability to convert other normally folded proteins of the same
kind to the misfolded form. The misfolded proteins have the tendency to
stack themselves in a tight arrangement called an aggregate. In humans, the
Prpc protein is located in the brain. The misfolding of Prpc forms large
aggregates within the brain and is associated with CJD.
[Het-s] is a prion found in the fungus of Podospora anserina that is caused by the Hets protein. During prion formation, the Het-s protein forms ribbon-like structures
(Fig. 4). These ribbons then give rise to small aggregates in the cell, which are
associated with the [Het-s] prion. Currently, the [Het-s] prion is of interest because it
is thought to have a structure similar to that of the human Prp prion.
Prions in History
In the past, infectious diseases have been associated with fungi, parasites, viruses,
and bacteria. In 1982, Dr. Stanley B. Prusiner suggested that prion diseases were
caused by aggregating infectious proteins and won the Nobel Prize in 1997 for his
work1. Prion proteins are associated with Creuzfeldt-Jakob disease (CJD) in humans
and bovine spongiform encephalopathy (BSE), also known as “mad cow” disease.
Mad cow disease received wide publicity in the mid 1990’s after cattle were fed
bone meal from prion infected sheep. The diseased cattle were then thought to
have entered the food supply for humans, causing an outbreak of CJD. The public
outcry in response to the BSE and CJD outbreaks challenged practices on how
nations raise and monitor meat products. In the United States, comprehensive
changes resulted from a “mammalian-to-ruminant feed ban through its BSE
inspection and BSE feed testing programs2.”
Figure 1. Creutzfeldt-Jakob disease and
age adjusted death rate. The number of
people who have died from CJD each
year (blue bars) has increased between
1979 and 2008, although the rate of
death among the population remains at
approximately 1 in a million3.
Figure 3. Prion protein aggregation. A normally folded prion protein (circles)
undergoes misfolding (becomes a triangle). The misfolded protein is able to
switch a normally folded protein to the misfolded prion form (as shown by the
small arrows). The accumulation of misfolded proteins leads to the formation
of aggregates, which create sponge-like holes when deposited in the brain.
Prion Aggregates Made of Beta Sheets
Balguerie, A., Dos Reis, S., Coulary-Salin, B., Chaignepain, S., Sabourin, M., Schmitter, J.M., and Saupe, S.J. (2004)
The sequences appended to the amyloid core region of the HET-s prion protein determine higher-order aggregate
organization in vivo. J Cell Sci. 117, 2610 .
Figure 5. The [Het-s] forms ribbonlike structures during prion
formation5. When Het-s protein
fuses to the Green Fluorescent
Protein, long ribbon-like structures
are formed during the initial stages
of [Het-s] prion formation (bottom
panel). These ribbons later give
rise to fluorescent aggregates.
Structure of the [Het-s] Prion
Crystal structures of the [Het-s] prion indicate that the prion has a solenoid
structure, as described in Fig. 4, in which the beta sheets are “stacked”.
Hydrophobic (V244, L276, F286, W287) and hydrophilic (Q240, E280) amino acids
are thought to keep this spring-like structure stable.
The conversion of the normal protein to the misfolded form results in a
structural change. The misfolded protein is rich in beta sheets. This
secondary feature is thought to be necessary for the formation of the
aggregates. It has been proposed that the newly misfolded protein uses the
beta sheet conformation to assemble into fibril like aggregates.
Image modified from
http://eebweb.arizona.edu/faculty/ma
sel/research/prionreplication/index.html
Figure 4. Beta sheets stack into a solenoid. Using the beta sheets, the
misfolded proteins stack together to create a solenoid structure, similar to a
coil in a spring. This solenoid structure generates the fibril appearance
common in prion aggregates.
A
B
C
Figure 6. Physical models of the [Het-s] prion based on 2KJ3.pdb. Physical protein
models were built using ZCorp 3D printing technology. Three different angles of the
models are shown. Hydrophobic amino acids (varying shades of blue) and
hydrophilic amino acids (varying shades of red) involved in solenoid stabilization can
be seen in A and C. The beta sheet “stacks” within the solenoid are visible in B.
"CJD (Creutzfeldt-Jakob Disease, Classic)." Centers for Disease Control and Prevention.
Centers for Disease Control and Prevention, 13 Dec. 2010. Web. 28 Feb. 2012.
Study of Fungal Prions
Prion Diseases Lead to Sponge-like Holes in Brain Morphology
Human prion aggregates are highly resistant to heat treatment and
detergents. Therefore, resistance to destructive techniques and potential for
infectivity make human prions possibly dangerous to work with in the
laboratory. Instead, prions found in organisms dissimilar to humans are used.
Prions found in fungus are also caused by an infectious protein and behave
similarly to their mammalian counterparts. Using fungal prions as a model
allows researchers to gain insight into the molecular structure of human
prions without risking infection. Researchers have focused on the particular
fungal prion [Het-s] of Podospora anserina .
Patients with CJD experience symptoms that include
dementia, poor motor coordination, and memory loss.
Progression of the disease is rapid and always fatal.
Figure 2. Normal versus classic CJD brain tissue. Cross
sections of post-mortem brains of CJD patients show large
holes in the tissue4. This spongy appearance is the reason
for its descriptive name, spongiform encephalopathy.
Genetic Science Learning Center. "The Mystery of
Kuru." Learn.Genetics 28 February 2012
A SMART Team project supported by the National Institutes of Health Science Education Partnership Award (NIH-SEPA 1R25RR022749) and an NIH CTSA Award (UL1RR031973).
Conclusion
Since the outbreak of bovine spongiform encephalopathy in cows and CreuzfeldtJakob disease in humans during the mid 1990’s, scientists have strived to
understand prions. Because of the infectious nature of the misfolded proteins,
researchers have looked into the fungal prion of [Het-s]. Human prions are thought
to form solenoid structures similar to [Het-s]. Therefore, studying the [Het-s] prion
structure could potentially lead scientists to new insights in relation to human
prions. The study of fungal prions provide a better understanding of human prions,
and potentially lead to the development of new medicines for prion-related
diseases.
References
1.
Prusiner, S.B. (1982). Science. 216, 136-144
2.
Bovine Spongiform Encephalopathy, or Mad Cow Disease, Centers for Disease Control and Prevention. http://www.cdc.gov/ncidod/dvrd/bse/
3.
Creutzfeldt-Jakob Disease. Centers for Disease Control and Prevention. Centers for Disease Control and Prevention
4.
The mystery of Kuru. Learn Genetics. Genetic Science Learning Center. University of Utah. http://learn.genetics.utah.edu/content/begin/dna/prions/kuru.html
5.
Balguerie, A., et al (2004). J Cell Sci. 117, 2610
6.
Van Melckebeke, H.et al (2010). J.Am.Chem.Soc. 132: 13765-13775