STUDY OF MOLECULAR DYNAMICS OF HALOPHILIC ?-AMYLASE ISOLATED FROM HALOTHERMOTHRIX ORENII AT DIFFERENT SALT CONCENTRATIONS

• Main Author: Eka Putri, Rahmaditya
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/49303
• Institution: Institut Teknologi Bandung

The enzymes isolated from halophilic organisms have a stable function and conformation at high salt concentrations. Thermostable and halostable friendly enzymes are required in industries involving processes with extreme conditions. One of the enzymes that has a very large application potential in the industry is AmyA, which is ?-amylase isolated from the halophyll bacterium Halothermothrix orenii. This enzyme has been known to have unique adaptability to environments with high salt concentrations and high temperatures. However, the research carried out so far has only been at the stage of purification and structural characterization of AmyA. Information about the reason of this enzyme has thermostable and halostable properties has not been widely explored. The purpose of this study was to store the stability of AmyA against salt levels and high temperatures through a molecular dynamic simulation approach. Mesostable enzyme (Amylase-G4) was used as a comparison in research. Molecular dynamics simulations were carried out for 10 ns at 500 K in a salt-free system and a 2 M NaCl concentration system. The results showed that the stability of AmyA could be maintained in a system without salt and a system with 2 M NaCl as seen from the gyration radius profile and nonpolar SASA. Although the stability of AmyA can be maintained, the system at 2 M NaCl concentration, experienced a nonpolar SASA decrease in the first 4 ns of the simulation. The decrease is adjusted for a regular increase towards the relatively similar nonpolar SASA values at the start of the simulation (0 ns). AmyA is known to have a different structure stabilizing strategy from common halophilic enzymes. The presence of flexible residues in AmyA, which is observed through the RMSF profile and molecular visualization, is a form of AmyA adaptation that maintains stability against high temperatures. Some of the AmyA residues with RMSF values that were striking in the unsalted conditions were Lys166, while those at 2 M NaCl conditions were Asn149 and Asn498 residues. AmyA does not have a protein surface which is dominated by negatively charged residues. The residue of AmyA contains a lot of hydrophobic residue and has a small residue. These adaptations make AmyA have good stability at high temperatures with no salt conditions. Overall, the high salt concentration does not really affect the stability of AmyA because it is halostable. In contrast to AmyA, Amylase-G4 has better thermal stability under 2 M NaCl conditions than without salt conditions. Another adaptation that helps Amylase-G4 to be able to maintain the stability of its tertiary structure is the high residual content of a small size so that it remains flexible at high temperatures. In addition, the presence of polar residues on the surface of the protein helps hydration at high salt concentrations. Some of the Amylase-G4 residues with RMSF values that were striking in the saltless system were Ser72 and Gln301, while the most flexible Amylase-G4 residues were Ala131, Gly300, and Ser399 in the 2 M NaCl solution. This research is able to provide detailed information on the atomic scale regarding the different adaptation mechanisms of AmyA and Amylase-G4 at high salt concentrations