Optimization of expression and purification of mitochondrial HSP 40 (Tid1-L) chaperone: Role of mortalin and tid1 in the reactivation and amyloid inhibition of proteins

Saudi Journal of Biological Sciences

Anwar Ahmed¹  Ajamaluddin Malik²  Wajhul Qamar²  Mohd Shahnawaz Khan²  Mohammad Rashid Khan³ 

[1] Corresponding author.;Department of Biochemistry, College of Sciences, King Saud University, Riyadh, Saudi Arabia;Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, Saudi Arabia;

关键词: Chaperones;    Refolding;    Amyloid;    Mortalin;    Insulin;   

DOI  :  

来源: DOAJ

【 摘 要 】

Stimulation of complex chaperone activity may be a viable means of therapy for neurodegenerative diseases. These chaperons execute reactivation of thermally and chemically aggregated protein substrates by cooperating with their partner co-chaperons. We optimized the expression and purification conditions of Tid1-L chaperone. Expression of Tid1-L in E. coli resulted in the formation of inclusion bodies which was further purified to soluble active form using 8 M urea and Ni-NTA column. Also, we investigated the events of the reactivation and disaggregation using aggregated G6PDH, luciferase and insulin as substrates. Incubation of aggregated/denatured enzymes with mortalin but not with Tid1 and/or Mge1 resulted in the initiation of the disaggregation reaction albeit very insignificantly. Under the same conditions coincubating the samples with chaperon and its assisted partners Tid1-L and nucleotide exchange factor Mge1 led to (40%) increase in enzyme activity of G6PDH. Similarly, luciferase activity was synergistically enhanced in the presence of mortlain/Tid1-L/Mge1 chaperones machinery. Chaperone-dependent disaggregation of thermally aggregated insulin showed that addition of Hsp70 and Hsp40 chaperones resulted in fast-track of renaissance reaction and inhibition of amyloid. The present study results conclude the quality of cell-control involves interaction of chaperon Hsp70 and its co-chaperones leading to complex formation with chemically/thermally aggregated substrate eventually causing its reactivation and disaggregation.

【 授权许可】

Unknown