Jake Zelie

BIOL 293
Questions- Studying a Prion
-synuclein (aS) is a neurological, infectious protein that generally forms
aggregates within patients that have Parkinsons disease. Proteins that are infectious are
generally referred to as prions, which are created due to misfolding in proteins. Prions
are not an exclusive phenomenon of mammals; they also occur naturally in unicellular
organisms such as yeast, which therefore have become good experimental models for
studying these protein conformational changes.(Nature) Accumulation of these prions
can cause an amyloid disease, like Parkinsons, and the accumulation of these proteins
can happen sporadically or genetically. Research shows, that as humans increase in age,
the chances for having a disease like Parkinsons increases dramatically. This is due to
the fact that the aging process increases mutations which interrupt the synthesis, folding,
and degradation of proteins, and this leads to the formation of misfolded protein
aggregates like aS. Alpha-synuclein is believed to function in synaptic vesicle
trafficking, and a portion of aS will form aggregates called Lewy bodies during aging
which is how Parkinsons forms. Alpha-synuclein also forms physiological multimers
that have the secondary protein structure known as the alpha helix conformation. This
prion is important, because researchers have found that alpha-synuclein can transmit
multiple system atrophy (MSA) which in turn causes health concerns for medical
professionals that operate on the infected brains of patients with these neurodegenerative
diseases. Proteins cannot be killed and this draws additional concern because
standard disinfection techniques that kill microbes do not eliminate the PrP prions that
cause CJD.(Science Daily)

Based on the image above from page two, it can be seen that this fragment of amino acid
residues has a polar charged amino terminal and a nonpolar uncharged carboxyl terminal. The
overall structure contains two positively charged R-groups, one negatively charged R-group, and
three uncharged R-groups. This means that the fragment has an overall charge of +1 which
makes the fragment quite hydrophobic. I believe that this fragment would most likely be found
in the core of the protein due to its hydrophobic nature.

In terms of structure both the normal and pathological forms of the alpha-synuclein protein
have alpha helices for secondary structure. The tertiary structure for the alpha-synuclein is
unknown for the pathological and normal forms. The quaternary structure of the alpha-synuclein
is not present for the pathological and normal forms. Based on the significance statement, the
authors intend to inform the public about the alpha-synuclein protein and also to explain how it
causes Parkinsons disease. The authors also intend to inform us that by substituting different
amino acid residues into the original alpha-synuclein strand KTKEGV, it can destroy
tetrameric forms and create alpha-synculein derivatives that do not form protein aggregates.

According to figure 2A, KTKEGV is the reference sequence; however KTKEGV only
appears fully in two samples. However if you look at positions one through six, you will notice
that K shows up seven times in the first position, T shows up six times in the second position, K
shows up five times in the third position, E shows up six times in the fourth position, G shows up
seven times in the fifth position, and V shows up six times in the final position. So based on
these values, these amino acids are the most common at the various positions they are in, which
means that the reference sequence is correct.
(Table used for questions 7-9)

For mutation KTKKGV, the change in amino acid was from a polar negative R
group to a polar positive R group, and it did not form tetramers. KTEEGV had an amino

acid change of polar positive to polar negative, and it did form tetramers. KTKEIV
changed from a small amino acid group to a large nonpolar group, and it did not form
tetramers. GTKEGV had an amino acid change of polar positive to small and nonpolar
and formed tetramers. KLKEGV had an amino acid change of a polar uncharged group
to a polar positive charged group and did not form tetramers. KTKEGR changed from a
nonpolar amino acid group to a polar positive charged group and formed tetramers.
KTKEGW had an amino acid change from nonpolar to aromatic and did not form
tetramers. In the panel A blot I can see two large bands at the alpha S14 position, and a
clear band at the alpha S60 position. In the panel B blot, I can see a band at the alpha
S14 position. In the panel C blot, I can see a clear band at the alpha S60 position. In
the panel D blot, I can see a band at the alpha S60 position.
Based upon the bar graphs, mutations KTKEGV, KTKKGV, KTKEIV, KLKEGV,
KTKEGW are all cytotoxic. Mutations KTEEGV, GTKEGV, and KTKEGR were not
cytotoxic. Mutations KTKEGV, KTKKGV, KTKEIV, KLKEGV, and KTKEGW formed
aggregates in human neurons and in rat neurons. Mutations KTEEGV, GTKEGV, and
KTKEGR did not form aggregates in human neurons and rat neurons. Aggregates are
important, because if numerous aggregates are formed in the body, you will be at higher
risk for neurodegenerative diseases such as Parkinsons disease.
The pattern that I noticed with the data analysis is that only mutations KTEEGV,
GTKEGV, and KTKEGR were not cytotoxic, and did not form aggregates in human
neurons and rat neurons. This tells me that if the peptide amino acid sequence is changed
into any of those mutations, then aggregates will not form. Based on the data analysis
and the significance portion of the article, the authors statement that amino acid
substitutions of the KTKEGV strand result in mutations that do not form aggregates is
correct.
After reading this article, I realized that healthcare and medicine can be
revolutionized if more time and analysis is devoted into the study of infectious prion
proteins. This article showed that if an amino acid strand is manipulated in such a way
that substituting another amino acid group can result in a strand that does not form
aggregates and is not cytotoxic, then there can be a way to cure such neurodegenerative
diseases such as Parkinsons disease or Huntingtons disease or CJD. This could expand
the medical world past such boundaries such as Alzheimers disease, which also has no
cure. I also realized that since you can't kill a protein,"(Science Daily) that prions are
among the worst infectious beings that exist in our world. However, if we can find a way
to prevent proteins from misfolding then prions will not be able to form, and aggregates
will not be able to form as well. Eventually, medical professionals should be able to find
a way to isolate proteins with repetitive amino acid motifs, such as polyglutamine,
(Nature) then certain neurodegenerative diseases will cease to exist.

CITED SOURCES:
University of California - San Francisco. "New type of prion may cause, transmit
neurodegeneration: Multiple System Atrophy is described as first new human prion disease
identified in 50 years." ScienceDaily. ScienceDaily, 31 August 2015.
Reynaud, Enrique. "Protein Misfolding and Degenerative Diseases." Nature.com. Nature
Publishing Group, 2010. Web. 6 Dec. 2015.