Jentashapir J Health Res. 2017 August; 8(4):e64461. Published online 2017 August 20. doi: 10.5812/jjhr.64461. Research Article Protective or Deteriorative Effect of Zinc Ions on Protein Misfolding: A New Insight into Amyloidogenic Disease Sepideh Noorzadeh, 1 Mohammad Reza Dayer, 2,* and Saeed Aminzadeh 3 1 Nourdanesh Institute of Higher Education of Meymeh, Esfahan, IR Iran 2 Department of Biology, Faculty of Science, Shahid ChamranUniversity, Ahvaz, IR Iran 3 National Institute of Genetic Engineering and Biotechnology, Shahrak-e Pajoohesh, Tehran, IR Iran * Corresponding author: Mohammad Reza Dayer, Department of Biology, Faculty of Science, Shahid Chamran University, Ahvaz, IR Iran. Tel: +98-6133331045, Fax: +98-6133331045, E-mail: mrdayer@scu.ac.ir Received 2017 March 15; Revised 2017 September 26; Accepted 2017 October 10. Abstract Insulin as a small molecule with 51 residues is an interesting model useful in studying protein misfolding of neurodegenerative amyloid proteins. Investigating zinc effects on insulin misfolding and aggregation triggered by 80% ethanol is the main objective of the present work. Using different methods of turbidity measurement, examining thioflavin T fluorescence changes during insulin aggregation, and conducting Far-UV circular dichroism spectroscopy of the process, we studied insulin aggregations in the presence of micromolar and millimolar concentrations of zinc ions to shed light on the mechanism of misfolding in these circumstances. Our findings confirmed that millimolar concentrations of zinc protect insulin integrity against acidic pH, high temperature, and 80% concentration of ethanol as misfolding inducer. We hypothesize zinc to be of therapeutic importance in amyloidogenic disease in case it is applicable. Keywords: Zinc, Insulin, Amyloidogenic Proteins 1. Background Neurodegenerative disorders, Alzheimer, Parkinson, and type II diabetes mellitus, are caused by the formation and extracellular deposition of fibrillar amyloids for cer- tain proteins (1-3). There are increasing reports indicating the involvement of biometals, e.g. zinc and copper, in this process (4-6). The fibrillation process consists of three dis- tinct steps: nucleation, elongation, and saturation (or pre- cipitation) of amyloid fibrils. Kinetic studies of protein fib- rillation show that in the first step of fibrillation, a prelimi- nary nucleus of the partially unfolded monomer is derived from the native protein (7). This step expected to be facil- itated by high temperatures, high concentrations of pro- tein, acidic pH, vigorous agitation, detergents, and organic solvents exposure (8). In the next elongation step of fib- rillation, a protofibril structure is formed by the associa- tion of the nucleus. In the last step, protofibrils are assem- bled to form highly ordered and thermodynamically sta- ble structures rich in beta sheets with least water solubility (9, 10). The precise mechanism of nucleation underlying the pathogenesis of amyloidogenic diseases remains to be characterized. Insulin is a small protein with 51 residues that make a reliable model to simulate protein misfolding experimen- tally. In acidic pH, high temperature, agitation, and/or in presence of organic co-solvent, insulin forms fibrillar structures in amyloidogenic diseases (11). Insulin is composed of two chains, A-chain with 21 residues and B-chain with 30 residues. These two chains are cross-linked by two interchain disulfide bonds (CysA7 - CysB7 and CysA20 - CysB19). There is also a third intrachain disulfide bond bridge made between CysA6 and CysA11. In- sulin in its secondary structure contains about 50% alpha- helical structures (including segments of A1 - A8, A12 - A19, and B9 - B19) and about 10% beta structures (segment B24 - B28). Two beta turns (residues B6-B9 and B20-B23) com- prise 16% of insulin structure and there is about 24% esti- mated random coil structure with about 15 residues (7-12). Ethanol as an organic solvent diminishes hydrogen bonds between water and protein backbone that decreases protein stability and pushes it toward denaturation and misfolding (13). There are some reports showing that low concentra- tions of ethanol exert a preservative effect on beta-amyloid protein and prevent aggregation. However, there is in- creasing evidence showing that high concentrations (> 80%) of ethanol trigger protein aggregation especially at Copyright © 2017, Jentashapir Journal of Health Research. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (http://creativecommons.org/licenses/by-nc/4.0/) which permits copy and redistribute the material just in noncommercial usages, provided the original work is properly cited