Acta Scientific Microbiology (ISSN: 2581-3226)

Research Article Volume 5 Issue 4

Expression and Characterization of in vitro Aggregates and Inclusion Bodies of α-synuclein in E. coli

Sudeepa Srichandan1, Kripa N Nand2* , Amulya K Panda1* and Pratima Ray3

1Product Development Cell, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India

2Department of Biological Sciences, RPI, New York, USA

3Faculty of Science, Jamia Hamdard University, New Delhi, India

*Corresponding Author:Amulya K Panda and Kripa N Nand, Product Development Cell, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India and Department of Biological Sciences, RPI, New York, USA.

Received: March 03, 2022; Published: March 24, 2022

Abstract

α-synuclein (SNCA gene) aggregation in neuronal cells causes Parkinson’s disease (PD), one of the most common neurodegenerative diseases. Mutations in the SNCA gene locus and accumulation of metal ions are the major hallmarks in PD patients. Inclusion bodies of α-synuclein were expressed in E. coli by incorporating mutations in α-synuclein and supplementing metal ions in the culture media. The inclusion body aggregates were purified and analyzed for the presence of amyloid type structures using thioflavin binding. The structure and functional characteristics of inclusion body aggregates formed during protein expression in E. coli makes them a suitable model to understand the mechanism of amyloid formation. Soluble α-synuclein was aggregated to form amyloid type aggregates in vitro. Inclusion bodies of α-synuclein showed similar structure and similar kinetics of formation as that of in vitro aggregates of α-synuclein. Formation of α-synuclein as inclusion bodies in E. coli and its amyloidogenic characteristics can be used to understand the process of protein aggregation. This information will be useful in discovery of next generation inhibitors for Parkinson’s disease.

Keywords: α-synuclein; Aggregation; Inclusion Body; Amyloids; Parkinson’s Disease

References

  1. A Ramón., et al. "Inclusion bodies: not that bad". Frontiers in Microbiology56 (2014).
  2. A Surguchev., et al. "Conformational diseases: Looking into the eyes". Brain Research Bulletin 81 (2010): 12-24.
  3. TF Outeiro., et al. "Dementia with Lewy bodies: an update and outlook". Molecular Neurodegeneration5 (2019).
  4. MG Spillantini., et al. "α-Synuclein in Lewy bodies". Nature 388 (1997): 839-840.
  5. B Dehay., et al. "Targeting α-synuclein for treating Parkinson’s disease: mechanistic and therapeutic considerations". Lancet Neurology 14 (2015): 855-866.
  6. MC Chartier-Harlin, et al. "α-synuclein locus duplication as a cause of familial Parkinson’s disease". Lancet 364 (2004): 1167-1169.
  7. Polymeropoulos MH., et al. "Mutation in the alpha-synuclein gene identified in families with Parkinson’s disease". Science80 (1997): 2045-2047.
  8. CM Lill., et al. "Genetics of Parkinson’s disease". Molecular and Cellular Probes 30 (2016): 386-396.
  9. B Fauvet., et al. "α-Synuclein in central nervous system and from erythrocytes, mammalian cells, and Escherichia coli exists predominantly as disordered monomer". Journal of Biological Chemistry 287 (2012): 15345-15364.
  10. L Pieri., et al. "Fibrillar α-synuclein and huntingtin exon 1 assemblies are toxic to the cells". Biophysics Journal 102 (2012): 2894-2905.
  11. L Pieri., et al. "Structural and functional properties of prefibrillar α-synuclein oligomers". Scientific Reports 6 (2016): 24526.
  12. M Ingelsson., et al. "Alpha-synuclein oligomers—Neurotoxic molecules in Parkinson’s disease and other lewy body disorders". Frontiers in Neuroscience 10 (2016): 408.
  13. ER Dorsey., et al. "The Parkinson pandemic—A call to action". JAMA Neurology 75 (2018): 9-10.
  14. PL Eleonora Carboni., et al. "Insights on the interaction of alpha-synuclein and metals in the pathophysiology of Parkinson’s disease". Metallomics 7 (2015): 395-404.
  15. L Mezzaroba., et al. "The role of zinc, copper, manganese and iron in neurodegenerative diseases". Neurotoxicology 74 (2019): 230-241.
  16. K Acevedo., et al. "Redox active metals in neurodegenerative diseases". Journal of Biological Inorganic Chemistry 24 (2019): 1141-1157.
  17. , et al. "Increased oxidative stress exacerbates α-synuclein aggregation in vivo". Journal of Neuropathology and Experimental Neurology 77 (2018): 443-453.
  18. AL Friedlich., et al. "The 5′-untranslated region of Parkinson’s disease α-synuclein messenger RNA contains a predicted iron responsive element". Molceular Psychiatry 12 (2007): 222-223.
  19. B Chen., et al. "Interactions between iron and α-synuclein pathology in Parkinson’s disease". Free Radical Biology and Medicine 141 (2019): 253-260.
  20. F Molina-Holgado., et al. "Metals ions and neurodegeneration". BioMetals 20 (2007): 639-654.
  21. P Lingor., et al. "Alpha-synuclein and iron: two keys unlocking Parkinson’s disease". Journal of Neural Transmission 124 (2017): 973-981.
  22. FA Marston., et al. "The purification of eukaryotic polypeptides synthesized in Escherichia coli". Biochemical Journal 240 (1986): 1-12.
  23. NS de Groot., et al. "Amyloids in bacterial inclusion bodies". Trends in Biochemical Sciences 34 (2009): 408-416.
  24. Cano-Garrido., et al. "Supramolecular organization of protein-releasing functional amyloids solved in bacterial inclusion bodies". Acta Biomaterial 9 (2013): 6134-6142.
  25. MM Carrió., et al. "Construction and deconstruction of bacterial inclusion bodies". Journal of Biotechnology 96 (2013): 3-12.
  26. E Gifre-Renom., et al. "The biological potential hidden in inclusion bodies". Pharm 12 (2020): 157.
  27. P Singhvi., et al. "Bacterial Inclusion Bodies: A Treasure Trove of Bioactive Proteins". Trends in Biotechnology 38 (2020): 474-486.
  28. A de Marco., et al. "Bacterial inclusion bodies are industrially exploitable amyloids". FEMS Microbiology Review 43 (2019): 53-72.
  29. VN Uversky., et al. "Metal-triggered structural transformations, aggregation, and fibrillation of human α-synuclein: a possible molecular link between parkinson′s disease and heavy metal exposure". Journal of Biological Chemistry 276 (2001): 44284-44296.
  30. P Davies., et al. "Alpha-synuclein is a cellular ferrireductase". PLoS One 6 (2011): e15814-e15814.
  31. Q He., et al. "Alpha-synuclein aggregation is involved in the toxicity induced by ferric iron to SK-N-SH neuroblastoma cells". Journal of Neural Transmission 118 (2011): 397-406.
  32. MP Jackson., et al. "Why are functional amyloids non‐toxic in humans?". Biomolecules 7 (2017): 71.
  33. S Ventura., et al. "Screening protein aggregation in cells using fluorescent labels coupled to flow cytometry". Protein Misfolding Dis. Methods Protoc., Springer, New York, (2019): 195-212.
  34. Zhu ZJ., et al. "Differential interaction between iron and mutant alpha-synuclein causes distinctive Parkinsonian phenotypes in Drosophila". Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease 4 (2016): 518-525.
  35. Ostrerova-Golts., et al. "The A53T α-Synuclein mutation increases iron-dependent aggregation and toxicity". The Journal of Neuroscience 20.16 (2000): 6048-6054.
  36. Peng Y., et al. "Binding of alpha-synuclein with Fe (III) and with Fe (II) and biological implications of the resultant complexes". Journal of Inorganic Biochemistry 4 (2010): 365-370.
  37. Joppe K., et al. "The contribution of iron to protein aggregation disorders in the central nervous system". Frontiers in Neuroscience (2019).
  38. E Tan., et al. "Iron regulatory protein (IRP)-iron responsive element (IRE) signaling pathway in human neurodegenerative diseases". Molecular Neurodegeneration75 (2017).

Citation

Citation: Amulya K Panda., et al. “Expression and Characterization of in vitro Aggregates and Inclusion Bodies of α-synuclein in E. coli". Acta Scientific Microbiology 5.4 (2022): 134-145.

Copyright

Copyright: © 2022 Amulya K Panda., et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.




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