Thermostability improvement of enzymes used industrially or commercially would develop their capacity and commercial potential due to increased enzymatic competence and cost-effectiveness. Several stabilizing factors have been suggested to be the base of thermal stability, like proline replacements, disulfide bonds, surface loop truncation and ionic pair networks creation. This research evaluated the mechanism of increasing the rigidity of organophosphorus hydrolase enzyme by flexible loop truncation. Bioinformatics analysis revealed that the mutated protein retains its stability after loop truncation (five amino acids deleted). The thermostability of the wild-type (OPH-wt) and mutated (OPH-D5) enzymes were investigated by half-life, ΔGi, and fluorescence and far-UV CD analysis. Results demonstrated an increase half-life and ΔGi in OPH-D5 compared to OPH-wt. These results were confirmed by extrinsic fluorescence and circular dichroism (CD) spectrometry experiments, therefore, as rigidity increased in OPHD5 after loop truncation, half-life and ΔGi also increased. Based on these findings, a strong case is presented for thermostability improvement of OPH enzyme by flexible loop truncation after bioinformatics analysis.
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The authors would like to thank the manager and colleagues of Applied Biotechnology Research Centre, Baqiyatallah University of Medical Sciences, Tehran, Iran, for their useful discussions and critical reading of the manuscript. It should be noted that this paper was extracted from dissertation of the research project of Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Corresponding editor: BJ Rao
Communicated by BJ RAO.
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Farnoosh, G., Khajeh, K., Mohammadi, M. et al. Catalytic and structural effects of flexible loop deletion in organophosphorus hydrolase enzyme: A thermostability improvement mechanism. J Biosci 45, 54 (2020). https://doi.org/10.1007/s12038-020-00026-5
- Bioinformatics design
- loop deletion
- organophosphorus compounds
- organophosphorus hydrolase
- thermostability improvement