Celine Cheung
Senior Thesis | 2025
Properties of Alpha-Synuclein and Its Effects on Parkinson’s Disease
Celine Cheung
Introduction
Parkinson’s Disease is a neurological disorder that worsens over time. It affects movements and causes shaking, imbalance, and incoordination. It also causes ambulatory and communication problems as the disease continues. Though the reasons are currently unknown, it’s been found that Parkinson’s Disease affects people older than 60 years old and men more than women.1
Alpha-synuclein is a small protein encoded in the synuclein gene. It is mainly found in central nervous system neurons. However, its function in pathological and physiological situations is largely unknown. It is hypothesized that alpha-synuclein causes movement disorders, such as Parkinson’s Disease. Patients with Parkinson’s Disease lose a significant amount of dopaminergic neurons. The remaining neurons are Lewy Bodies, which are alpha synuclein containing groups. One of the clear symptoms of Parkinson’s Disease are the point mutations of alpha-synuclein and the multiplication of the synuclein gene. Increased amounts of alphasynuclein in the ventral tegmental area have been related to early Parkinson’s Disease–like symptoms.2
Alpha-synuclein contains 140 amino acids and is located mainly in the presynaptic terminals. It also contains amphipathic N-terminus, which is used in the lipid binding properties of alpha-synuclein. It
contains a non-amyloid component (NAC), which is the aggregation domain, as well as the acidic C-terminus, which has the role of calcium binding and inhibiting protein aggregation. Some scientists have suggested that alpha-synuclein is used in the regulation of typical cellular function through interacting with binding proteins, as well as neurotransmitter release, vesicular trafficking and oxidative stress. Alpha-synuclein is linked to both familial and sporadic forms of Parkinson’s Disease. The autosomal dominant form of Parkinson’s Disease has also been linked to Nterminal missense point mutations of alpha-synuclein.2
Alpha-synuclein is not thoroughly understood in its functions but it has been found that it interacts with synaptic vesicles and controls the release of neurotransmitters. It is also known that alphasynuclein is bound by membranes. But its structure is largely unknown. In order to learn more about this structure, an Electron Paramagnetic Resonance-based (EPR)and site-directed spin labeling method was created. It specifically analyzes alphasynucleins that are bound to lipid bilayers. Both pulsed EPR and continuous-wave EPR are used. Continuous-wave EPR is able to figure out the correct secondary structure and the lipid-exposed residues’ membrane immersion depth. Pulsed EPR is able to figure out the long-range distances. Through these methods, as well as molecular dynamic simulations, alpha-synuclein has been found to be an alpha-helical structure that is both curved and extended. Alpha-synuclein’s helix contains a superhelical twist that resembles right-handed coiled coils because they both have 11-aa repeats. Glutamic acid and Lysine residues are able to interact with zwitterionic headgroups because of the way that the alphasynuclein helix lies parallel to the curved membrane. This form of
alpha-synuclein is very different from the alpha-synuclein that is found in SDS, which is used commonly and is a membranemimetic detergent. The alpha-synuclein found in the detergent is made up of two antiparallel helices. This difference displays the value in studying alpha-synuclein’s structure specifically in a bilayer environment. This interaction between alpha-synuclein and membranes is very important both in its function physiologically and its aggregation that contributes to Parkinson’s Disease. The interaction between alpha-synuclein and synaptic vesicles likely controls the release of neurotransmitters and the size of the presynaptic pool. It is also thought that alpha-synuclein favors interacting with curved vesicles that have a similar size to synaptic vesicles. This reveals how important it is to study alpha-synuclein and membrane interactions and how it affects the functions of alpha-synuclein. Alpha-synuclein sometimes contains an alphahelical structure in the N-terminal region which is caused by interactions with negatively charged vesicles. The structure of alpha-synuclein has been debated over. Some believe that a membrane-bound alpha-synuclein takes a shape similar to alphasynuclein bound to SDS detergent micelle, which is made up of two antiparallel helices wrapped around a micelle. Others believe that alpha-synuclein has an extended helical shape. Detergent micelles are useful for NMR structure analysis because they are small and can create high-resolution NMR analysis. However, this has been thought to affect the structure of alpha-synuclein, or the protein analyzed by NMR. As such, it is hard to determine if the structure found in the NMR analysis with micelles bound to it is the same structure of the protein that is membrane-bound. It is possible for alpha-synuclein that its structure when it is in a
membrane containing environment is an extended helical structure and the SDS micelles that attach to it break apart these helices in NMR analysis. As such, it’s crucial to create a method to precisely determine membrane protein structure when it is in a lipid bilayer containing environment.3
Alpha-synuclein containing animal models were created in order to examine the function of alpha-synuclein. The animals with alpha synuclein knockouts, through the deletion of the first two exons, were able to take in and release dopamine in response to stimulus. However, there was a reduction in striatal dopamine. This evidence further backs up the claim that alpha-synuclein is related to the regulation of dopamine neurotransmission.1
Another study found that the removal of alpha-synuclein resulted in a decrease in dopaminergic neurons in the substantia nigra, which further supports the claim that physiological levels of alphasynuclein affect the development and growth of neurons in the brain. Complete removal of the synuclein gene decreases the learning, working, and spatial memory.1
Two different models of alpha-synuclein overexpression have been used, transgenic animal models and viral-mediated alpha-synuclein overexpression. Increased levels of alpha-synuclein have been proposed to insult the dopaminergic system. An alpha-synuclein mouse model that contains a Thy1 promoter demonstrated overexpression of alpha-synuclein in regions of the brain implicated in Parkinson’s Disease. At 14 months old, the animals had a large decrease in dopamine levels, which suggests that before substantial nerve loss and motor deficits, there are early changes to the dopamine neurotransmission.1
Though all of these animal models suggest many things about the relationship between alpha-synuclein and Parkinson’s Disease, none of them contains all of the pathophysiological conditions associated with Parkinson’s Disease. As such, it is difficult to determine the specific cellular aspects of the dopamine system that alpha-synuclein overexpression disrupts. In response, scientists have looked at some prototypical cellular and molecular targets in neurodegeneration in an attempt to figure out how alpha-synuclein overexpression changes and stops neuronal activity before cell death.1
Alpha-synuclein is a significant part of amyloid fibrils that are within Lewy bodies, which are deposits among cells that pathologically signal the presence of a neurodegenerative disease, such as Parkinson’s Disease. Intrinsically, alpha-synuclein is a disordered protein and likely changes drastically in order to create amyloid fibrils. The mechanism that is thought to cause the disease is the process of aggregation between alpha-synuclein monomers into amyloid fibrils through the use of oligomeric intermediates. But the reason for the aggregation is not well-known. Through investigating pH and temperature on both mutant alpha-synuclein and wild-type synuclein using molecular dynamics simulations, the aggregation mechanism can be better understood. The folding between an unfolded protein into a somewhat folded intermediate could be the reason for both aggregation and fibrillation. An intermediate alpha-strand was present in the hydrophobic NAC region of alpha-synuclein that could then transition into an alphasheet and start the assembly process. The water environment around the intermediate was analyzed to figure out how it affected the alpha-strand. This investigation provided evidence for a quality
of alpha-synuclein aggregation and a possible neuroprotective plan that could both help heal Parkinson’s Disease and its symptoms.4
An alpha-strand is made up of amino acid residues that alternate between alpha right helical conformations and alpha left helical conformations. An alpha-sheet is made up of two close alphastrands that are bonded with bifurcated hydrogen bonds. Later residues possibly contain a similar alpha-strand conformation if they consist of an alpha right, alpha left, alpha right conformation or an alpha left alpha right alpha left conformation. The backbone amino groups all face the same way in an alpha-strand. On the reverse side of that strand, all the carbonyl groups face the different direction. This ends up making the strand have one positive charged side and one negative charged side. The NAC region of alpha-synuclein is amyloidogenic. The residues 71-82 are hydrophobic and very important in assembling filaments. When those residues are deleted, alpha-synuclein is not neurotoxic or able to aggregate anymore. Additionally, when these residues exist by themselves in simulations, the segment is able to polymerize itself and create toxic amyloid fibrils, as well as promote the formation of fibrils for full-length human alpha-synuclein. A reversible transition between alpha-synuclein and a partially folded intermediate was started through either the raise of temperature or the reduction in pH. The changes in a protein’s environment impacts both the hydrophobic qualities and its charge. The higher the temperature, the more of a stronger hydrophobic force is used to fold. The lower the pH, the more the net charge of the protein is minimized, which causes the intramolecular charge-charge repulsion to lessen and allows a collapse to an intermediate that’s partially folded. Amino-terminally alpha-synuclein that has been
acetylated takes on compact conformations when it undergoes physiological cell conditions. These conformations protect the aggregation-prone NAC region from being visible to the cytoplasm and therefore fights against aggregation. Additionally, the acetylation of N-terminals lowers the number of alpha-strand structures.4
Alpha-sheets are atypical secondary structures that are found in proteins. They are also known as pleated sheets or polar pleated sheets. It is formed by hydrogen bonds between consecutive alphastrands. Alpha-sheet residues are not limited to a certain area of dihedral angles, unlike alpha-helices and beta-sheets. Instead, it contains alternating dihedral angles in both the right-handed and left-handed helical areas of its Ramachandran plot. Though alphasheets are not usually found in natural proteins, it is present in many amyloid diseases and thought to have a major part in them. Alpha-sheets are thought to be an intermediate state during the forming of amyloid fibrils, which has been seen through MD simulations. In amyloid diseases, it is also thought to be a toxic conformer. Alpha-sheet structures have been seen in transthyretin, prion proteins, beta two-microglobulin and lysozyme. All of these proteins and peptides play a role in various amyloid diseases and protein misfolding instances. They also all change structures from being a mostly random coil or alpha-helix structure to being a betasheet structure that is usually found in amyloid fibrils. The structure for flipping the peptide plane has been proved to be responsible for clear conversion between alpha-sheets and betasheets. Many crystal structures and NMR structures in the Protein Data Bank also contain the alpha-strand conformation. Other studies have shown that experimental anti-alpha-sheet peptides like
to bind with toxic amyloid precursors, otherwise known as the intermediate alpha-sheet conformer, instead of binding with mature fibrils or non-aggregated, nontoxic species. As such, these peptides are able to control the aggregation of amyloid proteins and demonstrate that alpha-sheets and alpha-strands exist beyond the computational and theoretical spaces.4
Alpha-synuclein is linked to many neurodegenerative diseases, otherwise known as synucleinopathies. They are not just found in Parkinson’s Disease Lewy bodies. Other diseases it might be found in include Down’s syndrome, Parkinson’s Disease dementia and the Lewy body variant of Alzheimer’s Disease. Alpha-synuclein amyloids have been thought to be diverse in both structure and functionality. However, the reason for why the same protein aggregates into different fibril polymorphs is mostly unknown. To investigate the structure function of premature fibrils and helix mature fibrils, two alpha-synuclein aggregation intermediate polymorphs are demonstrated. Not only do polymorphs differ in structure, but they also differ in cellular activity. Helix mature fibrils have a compact core and its seeding potency is low. They can also quickly internalize and transfer between cells. Premature fibrils have less structure and cannot transfer between cells. However, they have abundant pathology of alpha-synuclein and triggers the creation of aggresomes, misfolded protein aggregates, in cells.4
In the aggregation pathway, the conformational heterogeneity may cause fibril polymorphs that contain clear prion-like behavior. Synucleinopathies are groups of neurodegenerative disorders that contain the presence of alpha-synuclein amyloid fibril composed
bodies. In Parkinson’s Disease, these bodies are Lewy Bodies and Lewy Neurites. In multiple system atrophy, the bodies are mostly in oligodendrocytes and are called glial cytoplasmic inclusions. The amyloid fibrils that are connected to neurodegenerative disorders have prion-like behavior and are transmissible. Alphasynuclein forms fibril polymorphs which is why there are different disease phenotypes.5
Polymorphism exists both in recombinant fibrils and fibrils that come from a diseased animal or patient’s brain. The differences in alpha-synuclein fibril polymorphs are mostly in fibril diameter and the number of twists and protofilaments. Some polymorphs have a common cross beta sheet structure but contain different packing and inter-protofilament interfaces. The aggregation process is stochastic and contains multiple inter-convertible forms of the same protein.5
Alpha-synuclein aggregation uses a heterogenous population of an intermediate species. It is thought that intermediate species are able to form alpha-synuclein fibril polymorphs with the same assembly environment. This hypothesis is tested by isolating aggregation intermediates and creating premature fibrils and helix-matured fibrils, two fibrillar polymorphs. The data supports the idea that the relative population and type of intermediate species could decide the characteristics of fibril polymorphs and the biological activities that go along with the pathological traits in synucleinopathies.5
Amyloid fibril formation starts with converting soluble monomers to highly ordered beta sheet rich aggregates. This usually follows a sigmoidal growth kinetics, made up of three phases. The first phase is the lag phase, which is the time needed for nuclei to grow and
proliferate into aggregates detectable in a solution. Next is the elongation phase, which is the phase in which fibrils grow and elongate and the rate the fibrils form at is the highest. The last phase is the stationary phase, which is the steady state phase. Both primary nucleation and fibril elongation are fundamentally required when amyloids form from a pure monomeric solution. Fibrils grow through adding monomers to the ends of already existing aggregates. The overall aggregation kinetics is largely impacted when processes modify the number of fibril ends. It is hypothesized that through primary nucleation, initial nuclei form within milliseconds and after elongation, secondary nucleation events dominate most of the lag phase.5
Though alpha-synuclein is a protein containing 140 residues, neither cysteine nor tryptophan is present. The first sixty residues make up the N-terminal region. The next 35 residues make up the central region. The last 45 residues make up the C-terminal region. The N-terminal region is an amphipathic region. The C-terminal region contains mainly residues that are acidic. The NAC region is the hydrophobic central region and it is able to fold into a betasheet that is important in both cytotoxicity and aggregation. The fibril core region of alpha-synuclein could likely be the residues 71-82, which overrides the previous suggestion that it was residues 30-110. In order for alpha-synuclein to form amyloid fibrils, the structure might change dramatically. There are lots of internal and external factors that can change the rate of alpha-synuclein aggregation that exists in experiments. These factors could be things like pH, mutations within the NAC region and metal ions. In part of the N-terminal, the region of residues 38 - 53 can form into a beta-hairpin and could possibly be the reason for alpha-synuclein
aggregation. But the role of alpha-strands and alpha-sheets during alpha-synuclein aggregation has not been studied yet. Only three NMR structures of a full-length alpha-synuclein exist. The two PDB codes 1XQ8 and 2KKW are alpha-synuclein structures that are bound by micelle. The PDB code 2N0A is also a full-length alpha-synuclein that is a human fibril which causes diseases. High temperatures have been seen in MD simulations to speed up the aggregation process without changing the overall structure and path. Hydrogen bonds link together water molecules which creates different kinds of hydrogen-bonded water rings. Alpha-synuclein is a protein that is naturally unfolded. It has been demonstrated through multiple studies that the way that alpha-synuclein fibrillogenesis occurs is similar to how beta-amyloid fibrillogenesis happens, in that it occurs through multiple nucleation-dependent polymerization steps. They also both need seeds that come from an ordered nucleus that then is followed by oligomers growing through the addition of more monomers. The seeding from the nucleus needs a primary nucleation that comes from the creation of a fibril nucleus made up of monomers. During the fibril elongation stage, the formed nucleus grows and creates mature fibrils, which then starts a generation of new aggregates through using secondary paths. These generations can be surfacecatalyzed nucleation or the fragments of fibrils and are dependent on the amount of already existing fibrils. The secondary pathways end up in the creation of a fibril nucleus that then dissipates into a mature fibril during the last step which then works in the polymerization process. The first step of primary nucleation is very slow, especially compared to the fast paced nucleation through secondary pathways. MD simulations of a wild type alpha-
synuclein monomer were created and its environment was low pH and neutral with a high temperature. These simulations led to the hypothesis for reasons of forming alpha-strands and the hypothesis of an aggregation mechanism for the role of alpha-strands and alpha-sheets in the formation of alpha-synuclein structures.4
In order to further investigate alpha-synuclein properties and the effects of different polymorphs, three full-length alpha–synuclein polymorphs were chosen from the Protein Database (PDB) to be computationally analyzed in a box of water. Through observing the interactions of the water molecules with each alpha-synuclein polymorph, different qualities, including alpha-strand presence and pH environments can be analyzed.
Methods
The GROMACS tutorial of Lysozyme in a Box of Water was used to analyze three full-length alpha-synuclein polymorphs (PDB files 1XQ8, 2N0A, and 2KKW). The simulation recreates alphasynuclein in a box of water that contains ions. The PDB text file is the first necessary file because it visualizes the protein using programs such as VMD. Crystal water can be replaced from structures that are bound loosely or do not contain an active-site water molecule. The PDB file cannot contain missing atoms within internal sequences or residues because otherwise the pdb2gmx program will fail. There also cannot be any arbitrary molecules that pdb2gmx is expected to create because they are not defined.6
Pdb2gmx creates three different useful files for the molecule. It first creates the molecule’s topology. Then it creates a file that shows the molecules' restraint on position. The last file contains the structure after it has been processed. The topology file is typically called the topol.top file. It defines the molecule in a simulation, which means that the atom types, charges, bonds, and angles are all described. In the simulation, a force field must be chosen because it determines the information that the topology will contain. The force field chosen for alpha-synuclein is the OPLSAA/L all-atom force field. All the hydrogen atoms in the file were ignored because the naming system in the PDB file was not the same naming system in the force field. All the information in the topology file is within the force field. It defines the molecules as well as describes the system.6
The solvent used the SPC/E water. The next step of the simulation was to create the box through defining the structure and fill it with
the solvent, which is water. To do so, the box was created by describing the dimensions and taking advantage of the solvate module in order to fill the box with the water. A cubic box was chosen to be the unit cell because it is a simple structure. Alphasynuclein was placed within the box at least 1.0 nm from all of the box’s edges. This guarantees that there is at least 2.0 nm between any of the periodic images of a protein. As such, there is not a cutoff scheme that will harm the protein.6
Water solvation configure is defined in the GROMACS system. Spc216.gro was chosen as the configuration for the water because it is an equilibrated solvent model. The solvate counts the water molecules and updates the topology as it adds them to accurately reflect the system. The GROMACS solvate command only updates the topology if the solvent is water, as a feature of GROMACS. At this point, the system is solvated and contains a charged protein. In real life, proteins cannot live at a net charge, meaning that adding ions to the system is crucial for the experiment.6
Genion is a tool that is used to add ions within GROMACS. It reads the topology and replaces water molecules for ions. The input of genion is a run file, which means that it’s produced by the GROMACS pre-processor as an extension of a .tpr file. The .tpr file has the definitions and expectations for all the atoms in the system. A molecular dynamics parameter file (.mdp), is necessary to create the .tpr file. Grompp, or the GROMACS pre-processor, is able to use the information in the .mdp file, namely the topology and coordinates, in order to create the .tpr file. Usually .mdp files are used in order to run MD simulations or to run a minimization of energy, but this time it was used to create a description of the
system atomically. The .mdp file that the GROMACS tutorial provided was chosen because it was a basic script that did not have complicated elements. Through using one genion command, multiple things occur: a state file is imputed, a .gro file is outputted, the topology is processed and updated with both the removal of water and the addition of ions, positive and negative ion names are clarified and genion knows to only add enough ions to neutralize the net charge of the protein. It does this last part by adding the exact number of ions to neutralize the protein. At this point, the solvated system has no charge and is fully assembled. However, before molecular dynamics begins, the system must undergo the process of energy minimization. This ensures that there are no inappropriate geometry or steric clashes. Like the process of adding ions, energy minimization also uses grompp to create the structure, topology and parameters and deposits it into a .tpr file. The topol.top file must be updated in order for genbox and genion to run properly. The output of using mdrun for energy minimization are four em files. There is a text log file in ASCII that describes the whole energy minimization process. The energy file and full-precision trajectory file are in binary. Finally, there is a file that contains the structure with the energy-minimized. To figure out if the energy minimization worked, there are two tests that can be done. The first is whether the maximum force, which the minim.mdp set the goal maximum force, is reasonable. The second one is whether the potential energy is negative. Through using the em.edr file, which is the binary energy file, a potential.xvg file can be created which can be used to create an energy minimization plot in a program called Xmgrace. Finally, the system is ready to start the dynamics process. The first step is to
make sure the ions and the solvent are both equilibrated around the protein. If this step is skipped, there is a risk that the dynamics are unrestrained and could cause the system to collapse.6
Though the solvent is optimized, it is only within itself and not fully optimized with the solute. As such, it is necessary for the solvent and the solute to be at the ideal recreation temperature. It is also important for the solvent to be oriented in the right position around the protein, or the solute. After setting the temperature to the correct one, through using kinetic energies, the density must now be set by applying pressure to the system. The posre.itp file from a previous step is useful now. It applies a position restraining force on all the atoms of the protein that are not hydrogens, as those are the heavy atoms. The atoms are able to move once they have overcome the big energy restraint. This is useful because it makes it easier for the solvent to be equilibrated around the protein since there are less protein structural changes with the atoms restrained. The coordinates when the restraint potential is zero is also known as the origin of the position restraints. These are passed into the system through providing a coordinate file in grompp.
Equilibration occurs in two stages. The first stage occurs in a NVT ensemble, which means that the Number of particles, Volume and Temperature are all constant. It’s also known as a canonical ensemble, or an isothermal-isochoric ensemble. The contents of the system determines how long it takes for the procedure to run. The temperature, however, should plateau at a desired temperature. Through calling grompp and mdrun, parameters were added that included not coupling the pressure, and starting the generation of velocity. Now the temperature of the system should be stabilized through the NVT step. The second stage of equilibration before
analyzing the protein is to stabilize the pressure, which in turn stabilizes the density of the system. This is done in a NPT ensemble, which means that there is a constant Number of particles, Pressure and Temperature. It is also called an isothermalisobaric ensemble and is the environment that most closely mimics an experimental environment. The checkpoint file from the previous NVT step must be included because it has the state variables that are necessary to continue the simulation. This ensures that the velocities that are produced in the NVT equilibration step are conserved. Finally, the system is equilibrated in both temperature and pressure, which means that the restraints on the heavy atoms can be released and the production MD can be run in order to collect data. It is necessary to include a checkpoint file here as well in order to use the pressure coupling information that is preserved. One function of MDrun is that it decides the number of processors for the PP and PME calculations to work. Now that the protein has been simulated, it is able to be analyzed. Trijconv is a processing tool that is used to make up for any periodicity in the system. Alpha-synuclein was centered and the system was used as the output. The structural stability was looked at by calculating the least-squares fit and the RMSD calculation. By plotting the data, it shows the RMSD relative to the structure that is present in the system that was both equilibrated and minimized. The radius of gyration of the protein shows the measure of how compact it is. If the radius is relatively steady, that indicates that the protein is stably folded. If the radius varies over time, that indicates that the protein unfolds. The xtc file that is one of the outputs of this simulation is crucial as it is one of the trajectory or topology inputs.6
Discussion
Through these simulations, many qualities about the polymorphs were found. 2KKW simulations have a lower occupancy of alphastrands than 2N0A simulations. This is likely due to the fact that the 2N0A system, which is a fibril state, creates more alpha-strand structures than 2KKW, which is a micelle-bound state. Alphastrands form faster in a low pH environment, compared to a neutral pH environment. This makes sense compared with the pH as a catalyst for the partial unfolding that amyloid proteins undergo to become alpha-sheets. The appearance of alpha-strands is a lot slower in 2KKW system simulations compared to 2N0A system simulations which means that is is likely that the fibril state, or 2N0A, is able to become an intermediate alpha-strand quicker than a micelle-bound conformation, or 2KKW. The nucleationdependent polymerization because that also implies that fibrildependent secondary pathways are much faster than when a nucleus is created through monomers, otherwise known as the primary nucleation process. Alpha-strand was present in simulations with a high temperature. An alpha-strand segment of residues 72-74, which are Gly and Val amino acids respectively, was present in MD simulations. Because this came from a computational experiment, it is necessary to check whether the structure exists in real life. Through searching PDB files, which come from in person experiments, there were alpha-strands that did occur in the PDB files as a whole, which also supports the alpha right alpha left alpha right conformation that occurred in the residue region 72-74 in the MD simulation. However, the alphastrand structure did not appear in the residue region 72-74 in any of the PDB files.
All of this knowledge about alpha-synuclein is important in order to figure out the design of an alpha-synuclein inhibitor. It makes it more difficult to understand alpha-synuclein because it does not contain a clearly defined structure. Additionally, the way that alpha-synuclein aggregates is also unclear. The interactions between the proteins, which are intermolecular, usually include surfaces that are shallow and do not have ligand-binding sites that are clearly defined. As such, creating inhibitors for alpha-synuclein that work as drugs and attempt to prevent and reverse amyloid fibrils is very challenging. Instead, the drug that is worked on as more effective is used as peptides. This can be through designing short peptides that are synthetic which can stop alpha-synuclein from aggregating. There are also breakers made up of beta-sheets which bind to beta sheet precursors and then separate them. The created peptides can be made fast and economically saved through using chemicals. They can also be changed so that the binding properties improve. Peptides that degrade are also considered to be less toxic than the organic molecules that degrade, which are also man-made. Alpha-strands in the NAC region usually form alphasheets in oligomers. Alpha-sheets work like nuclei and start and encourage alpha-synuclein to both aggregate and fibrillate. There were multiple peptides created to fight against alpha-sheets and were successfully able to stop amyloid fibrils from forming and therefore fight against amyloid fibril cytotoxicity. This approach is promising because it is neuroprotective and could possibly help alleviate Parkinson’s Disease and its symptoms. The water environment around proteins strongly impacts the proteins. Both in real life experiments, as well as computer simulations, have
demonstrated that water has a huge impact on the function, structure, dynamics and stability of a protein. The role of water in both folding and unfolding of a protein is not largely known. This also means that the hydrogen bonds are important to study within the protein structure. By looking at the water-ring network in the NAC region alpha-strand of alpha-synuclein, the topical water network is analyzed. Water molecules likely play a huge role in both the protein folding and unfolding. The somewhat folded intermediate has been shown by multiple experiments to be the cause of the formation of amyloid fibrils through alpha-synuclein. Other amyloid proteins, such as prion proteins, transthyretin and lysozyme have also shown alpha-stands, and therefore alphasheets, to be this intermediate, which has been shown in various experiments. The alpha-sheet and alpha-strand structures have not been reported previously for alpha-synuclein. However, there was an alpha-strand that was shown in the NAC region of the protein, specifically in residue 72-74. When the NAC region is separated from the other residues, amyloid fibrils are able to polymerize themselves and therefore create toxic amyloid fibrils. As such, it is very crucial that alpha-strand occurs in the NAC region of alphasynuclein. TWN analysis for an alpha-strand residue has shown that the alpha-strand exists depending on the water surrounding it. Through targeting the specific region of residues 72-74, amyloid fibrils can be stopped, which is necessary for many neurodegenerative diseases, especially Parkinson’s Disease.4
References
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(4) Balupuri, A., Choi, KE. & Kang, N.S. Computational insights into the role of α-strand/sheet in aggregation of αsynuclein. Sci Rep 9, 59 (2019).
https://doi.org/10.1038/s41598-018-37276-1
(5) Mehra, S.; Ahlawat, S.; Kumar, H.; Datta, D.; Navalkar, A.; Singh, N.; Patel, K.; Gadhe, L.; Kadu, P.; Kumar, R.; Jha, N. N.; Sakunthala, A.; Sawner, A. S.; Padinhateeri, R.; Udgaonkar, J. B.; Agarwal, V.; Maji, S. K. α-Synuclein Aggregation Intermediates Form Fibril Polymorphs with Distinct Prion-like Properties. Journal of Molecular Biology 2022, 434 (19), 167761. https://doi.org/10.1016/j.jmb.2022.167761.
(6) Lemkul, J. Lysozyme in Water. www.mdtutorials.com. http://www.mdtutorials.com/gmx/lysozyme/index.html.