Active Alpha-Synuclein Proteins in Research

For education only

This guide cannot diagnose a condition or recommend a personal treatment plan. Discuss medical questions with a qualified professional.

Alpha-synuclein proteins have been implicated in several neuropathological diseases.

Image Credit: Naeblys/Shutterstock.com

The de-regulation of the protein is thought to influence Parkinson’s disease, dementia, and other illnesses through the build-up of its intraneuronal aggregates, known as Lewy bodies.

Research using active alpha-synuclein proteins has proven invaluable in understanding more about the role of Lewy bodies in pathology, along with assisting in the development of new pharmaceutical drugs for the treatment of Parkinson’s disease, Lewy body dementia, and other disorders.

Alpha-Synuclein’s role in disease

The human heart, gut, muscles, and brain are all home to the alpha-synuclein (α-syn) protein. While its functions are not 100% clear, there is a vast body of evidence that has implicated it as being fundamental to numerous key processes, most notably it appears to be essential for dopamine regulation.

Studies have shown that the protein is responsible for managing the process of transporting neurochemicals between synapses, known as neurotransmission.

Dopamine plays a fundamental role in the initiation and development of Parkinson’s disease (PD), and research has shown that there is a link between alpha-synuclein and the disease that looks likely to have a genetic basis. In addition to this, it has been found that alpha-synuclein is involved in multiple system atrophy (MSA), dementia with Lewy bodies (DLB), and pure autonomic failure (PAF).

Along with Parkinson’s disease, these diseases are known as the synucleinopathies, due to their close link with alpha-synuclein. Evidence has uncovered that pathological deposition of these protein aggregates, or Lewy bodies (LBs), are linked with the synucleinopathies.

Unsurprisingly, much research has been conducted in this area to understand the cause and development of these disorders, as well as to explore potential new therapeutic avenues.

Lewy bodies are aggregates of alpha-synuclein

The alpha-synuclein protein aggregates in neuropathological diseases, such as Parkinson’s. The formation of these toxic clumps is characteristic of the illness, and they are associated with the related death of brain cells.

For this reason, the de-regulation of the alpha-synuclein protein is considered to be an underlying cause of the synucleinopathies. Tests to identify these protein clumps or Lewy bodies is essential to early, accurate diagnosis because often symptoms of certain synucleinopathies resemble those of Alzheimer's or even psychiatric illnesses.

Testing for Lewy bodies, the intraneuronal aggregates of the alpha-synuclein protein, is therefore essential to the continued research of Parkinson’s disease, Lewy body dementia, and other disorders.

Research techniques with active alpha-synuclein proteins

Experimentally, active alpha-synuclein proteins can be used to induce Lewy bodies so that they can be studied in specific, controlled scenarios. These kinds of studies can elucidate the behavior of Lewy bodies, and uncover more about their role in various diseases.

Methods have also been developed to detect endogenous active alpha-synuclein protein levels in a sample using a dye.

These techniques can be used to study samples both in vivo and in vitro, and they can look into not only the functions of Lewy bodies in pathology but can also assist in the development of new pharmaceutical drugs through testing their impact on Lewy bodies.

Fluorescence techniques are the most common method for identifying active alpha-synuclein proteins. Thioflavin T (ThT) is one of the most popular dyes used in this assay. The process involves the seeding of new alpha-synuclein aggregates from a pool of active alpha-synuclein monomers.

The fluorescent ThT dye is added to bind to the beta sheet-rich structures that are found in alpha-synuclein aggregates. The event of ThT binding with the structures in the aggregates causes a shift in the dye’s emission spectrum, turning it red and increasing its fluorescence intensity.

Past research has used this technique to identify the aggregation of these endogenous proteins in Alzheimer’s disease (AD), Parkinson’s disease, amyloidosis, and others. The methods can be constructed also in vitro to allow for the study of interactions in the absence of other influencing factors.

Modern studies mostly perform assays with Thioflavin T within fluorescence plate readers, which allow for multiple conditions to be explored at the same time. While it is an established technique that is frequently used for looking at active alpha-synuclein and its aggregates, the process is not without its limitations.

The method is known for its poor reproducibility, mostly due to the number of varying factors that affect protein aggregation. Currently, scientists are working on improving this method to increase the reproducibility of ThT assays. Recent developments of the method have involved adding a period of orbital shaking, as well as improving mixing with the addition of glass beads.

Future directions

Active alpha-synuclein proteins provide a robust method for looking at the Lewy bodies associated with numerous neuropathological diseases. They are currently being used to understand the impact of alpha-synuclein on these diseases, as well as to develop new treatment methods.

The most common method of investigating them is through fluorescence techniques that use Thioflavin T as a dye. While this method is reliable and well established, scientists are currently working on improving its reproducibility to enhance the strength and accuracy of the method.

Sources:

Li, X., Dong, C., Hoffmann, M., Garen, C., Cortez, L., Petersen, N. and Woodside, M. (2019). Early stages of aggregation of engineered α-synuclein monomers and oligomers in solution. Scientific Reports, 9(1). https://www.nature.com/articles/s41598-018-37584-6

Narkiewicz, J., Giachin, G. and Legname, G. (2014). In vitro aggregation assays for the characterization of α-synuclein prion-like properties. Prion, 8(1), pp.19-32. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6066150/

Wördehoff, M. and Hoyer, W. (2018). α-Synuclein Aggregation Monitored by Thioflavin T Fluorescence Assay. BIO-PROTOCOL, 8(14). https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6066150/

Helpful context for this guide

Source-derived material selected through this article’s indexed topics.

Related questions

01What are the Advantages of Affimers?

Affimers are small in size, stable at different temperature ranges, relatively easy to generate, and offer appealing opportunities to complement the shortcomings of antibodies. Their small size is thought to be one of the attributes making affimers more sensitive than antibodies when used in histochemical staining, as they can penetrate tissues better. Affimers are both commercially available and can be synthesized by individual laboratories using publicly available methods, thereby ensuring it is accessible to the entire scientific community. One key advantage of affimers, as opposed to antibodies or nanobodies, is the speed at which they can be screened and developed. The screening process for affimers involves three iterations of phage display and ELISA assays. This can be carried out in 12 days, which is significantly less than the time needed for antibody and nanobody animal inoculation times. Furthermore, the screening process by which binding reagents are identified is more efficient because the target proteins are screened in a folded state.

Source: www.news-medical.net ↗

02Why do so many of the compounds fail?

One of the main reasons why compounds fail is that they actually don't work in the patients they are tested in. We may have picked the wrong target since although we've actually created a molecule that, for example, inhibits the activity of the protein in the disease, it may actually turn out that over activity of that e protein isn't as important as we thought it was in that disease. The other reasons molecules fail now is primarily unexpected toxicity. You could take a molecule all the way through to phase 3 trials – the biggest trials in patients – and suddenly find that in your long-term, two-year carcinogenicity studies in animals, that you've got something that is quite cancer-provoking, so you have to stop development. Those are the two reasons: efficacy and toxicity. Somehow, we've got to find a way of trying to increase the chances of success or making the process less costly and leaner.

Source: www.news-medical.net ↗

03With emerging fields like AI and machine learning, how are these technologies being integrated into your research and drug discovery to accelerate the identification of potential drug candidates?

While we've initiated dialogues on integrating AI, it hasn't transformed our work yet. However, I see it as a significant tool, especially for handling and processing big data. AI can help bridge the expertise gap in understanding both bioinformatics and biological terms, aiding in the efficient mining of information.

Source: www.news-medical.net ↗

04What would be the impact on cancer patients if COTI-2 does prove to be effective in people with p53 mutant tumours?

Given the central role of p53 mutations in human cancers, COTI-2 could represent a breakthrough therapy for many cancer patients if clinical trials confirm its’ activity in people. If we consider a specific disease such as ovarian cancer, p53 mutations are found in more than 90% of these tumours. Preclinical animal experiments with a human ovarian cancer known to have a p53 mutation and be resistant to conventional chemotherapy demonstrated that treatment with COTI-2 as a single agent either completely halted tumour growth or lead to dramatic tumour regression depending on the dose. COTI-2 was associated with no observable toxicity in these experiments.

Source: www.news-medical.net ↗

05What knowledge has your research provided, specifically regarding Alzheimer's disease?

The main protein or peptide in Alzheimer's disease patients is something called beta-amyloid. It has a 42 amino acid chain, and accumulates in the plaque between nerve cells in Alzheimer's disease patients. Beta-amyloid has been found to epimerize. Certain amino acids epimerize in the beta-amyloid at specific locations, and it turns out those amino acids are mainly aspartic acid and serine. There was no good way to analyze for such aberrations in beta-amyloid until now, and we have found a number of techniques that not only detect it, but can point out exactly where, within the beta-amyloid chain, the aberration is occurring and if there are more aberrations in Alzheimer's patients than in normal people. Image Credit:Shutterstock/Tavarius

Source: www.news-medical.net ↗

About the author

Peptide Therapy Guide Editorial Team

Editorial team for Peptide Therapy Guide.

View all articles →

Alpha-Synuclein’s role in disease

Lewy bodies are aggregates of alpha-synuclein

Research techniques with active alpha-synuclein proteins

Future directions

Sources:

Further Reading

Related questions

Peptide Therapy Guide Editorial Team

What are Affimers?

BIOVECTRA and Revolution Biomanufacturing partner to deliver integrated mRNA Design and GMP production services

Transgenic carrots producing nisin protein show promise as natural food preservatives

Cationic peptides inhibit SARS-CoV-2 cell entry