1st International and 10th National Iranian Conference on Bioinformatics
Home Accepted Papers
Protein Engineering of Bacillus α-Amylase to Improve Thermostability and Low Water Activity: A bioinformatics approach
Paper ID : 1140-ICB10
Authors:
Babak Elyasi Far *1, Maysam Mard-Soltani2, Ladan Mafakher3, shiva Mohammadi4
1Department of Physiology and Pharmacology, School of Medicine, Dezful University of Medical Sciences, Dezful, Iran
2Department of Clinical Biochemistry, Faculty of Medical Sciences, Dezful University of Medical Sciences, Dezful, Iran
3Thalassemia & amp; Hemoglobinopathy Research center, Health research institute, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
4Department of Medical Biotechnology School of Medicine, Lorestan University of Medical Sciences, Khorramabad, Iran.
Abstract:
Background: α-Amylase produced by Bacillus licheniformis (BLA) is widely used in starch industries. Water is so crucial for α-Amylase activity; due to the low amount of available water in the starch industry, it can lead to denaturation of the enzyme. In this study, we attempt to reduce BLA aggregation and improve low water activity by protein engineering without reducing its activity by computational methods.
Material and Methods: As surface residues are responsible in aggregation and water activity, the surface amino acids were defined by PyMol software (PDB ID: 1VJS). Aggrescan webserver was used to define hotspot aggregation residues. The surface amino acids of the α-Amylase which did not locate in the active site of the enzyme and were hotspot aggregation residues, replaced with glutamic and aspartic acid as negative amino acids by Chimera software. The stability and solubility of engineered α-Amylase compared to native form were done by Protparam and Protein-sol, respectively. To analyze secondary protein Psipred webserver was applied. Moreover, molecular dynamic simulation was done by Gromacs package to assess the stability of the tertiary structure of mutant form compared to native BLA during the time.
Result and Discussion: According to the active site analysis of BLA, surface Amino acid Q340E, S356D, G474E, L318E, and S310E have neutral charges and are recognized as hot spots of aggregation that was changed to negative charge amino acids. Physicochemical properties analysis, protein solubility, and aggregation potential discovered that these properties of engineered enzyme improved from engineered mutant compared to native form. Also, secondary structure evaluation indicates that these mutations caused no change in protein structure which was proved by molecular dynamics simulation during time. So, these mutations could reduce BLA aggregation and improve low water activity although experimental analysis needs to prove it.
Keywords:
α-Amylase, protein engineering, low water activity, In Silico
Status : Paper Accepted (Poster Presentation)