Cloning, expression and purification of the BRICHOS domain from Bri2 and its anti-amyloidogenic effects

Kardan, Nasrin; Dabirmanesh, Bahareh; Khajeh, Khosro

Cloning, expression and purification of the BRICHOS domain from Bri2 and its anti-amyloidogenic effects

Document Type : Original Research

Authors

Nasrin Kardan

Bahareh Dabirmanesh

Khosro Khajeh

Tarbiat Modares University, Faculty of Biological Sciences, Department of Biochemistry, Tehran, Iran

Abstract

Protein deposition due to the process of accumulation inside or outside cells causes many neurological diseases such as Alzheimerchr('39')s, Huntingtonchr('39')s or Parkinsonchr('39')s seizures. Parkinsonchr('39')s disease is the second most common neurological disease after Alzheimerchr('39')s, in which patients develop disorders due to the accumulation of leprosy and the destruction of dopamine neurons. Alpha-synuclein protein contains 140 amino acids, the main protein known in lewy body accumulations. During the aggregation process, alpha-synuclein protein monomers bind together as oligomers and eventually become amyloid filaments. So far, there is no drug to stop or delay the progression of Parkinsonchr('39')s, but studies on the molecular mechanism of amyloid formation and the identification of inhibitors are increasing. For this purpose, in this study, the effect of BRICHOS domain resulting from BRI2, which can have various functions, including antimicrobial properties, on the process of alpha-synuclein accumulation as a model protein was investigated.The gene was first optimized and synthesized and then multiplied by PCR. The product was digested by enzymes Xho I and Nde1 and entered the expression vector pET28 a, which was transformed into E. coli bacteria.Finally, the peptide was purified by nickel chromatography. The alpha-synuclein gene was also expressed separately and purified.The anti-cumulative effect of BRICHOS domain on alpha-synuclein fibrillation was investigated using Toflavin T fluorescence method and TEM technique.

Keywords

Amyloid

neurodegenerative disorders

BRICHOS domain

chaperone

20.1001.1.23222115.1399.12.1.9.1

Subjects

Molecular biotechnology

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