Kestabilan Termal Asam Amino Enzim Lipase Bacillus subtilis Menggunakan Simulasi Dinamika Molekul

Abstract

Bacillus subtilis lipase A (BsL) is extensively studied in enzymatic production of biodiesel, production of detergents, surfactants, food, pharmaceutical products, cosmetics, paper industry and nutrition. One drawback of using BsL is its relatively low optimum temperature at 37 oC (310 K). The objective of this research is to design BsL mutant more thermostable than the wild-type BsL by engineering mutations in residue stabilizer BsL. Molecular dynamic simulation was conducted to get better insight on the process of thermal denaturation or unfolding in BsL. Thermal denaturation of BsL was accelerated by conducting simulation at high temperature. Molecular dynamic simulation of BsL was performed with NAMD software package at 400-500 K. Selection of mutated residues was based on electrostatic interaction, disulfida bond, hidrofobic interaction analysis of BsL. From those analyses, three mutants were designed, which are Mutant-1 (Iso155Val), Mutant-2 (Ile135Cys/Hsd156Cys), and Mutant-3 (Glu65Asp). Parameters that were used to compare the thermostability of mutant with wild type enzyme were RMSD (Root Mean Square Deviation), SASA (Solvent Accessible Surface Area), Rg (Radius of Gyration), RMSF (Root Mean Square Fluctuation) and Secondary Structure. Molecular dynamics simulations were performed on all three mutants showed that the mutant-1 (Iso155Val) by giving the temperature of 525 K for 20 ns has a better thermostability compared to the wild type BSL. It concluded that hydrophobic interaction is important in improving the thermal stability of the enzyme lipase B. subtilis. Effect of mutation of hydrophobic interaction analysis results in an increase in the thermal stability of mutant-1 (Iso151Val) by giving a temperature of 500 K for 50 ns can improve thermal stability as evidenced by a decrease in the value of the parameter RMSD, Rg, RMSF, SASA and Secondary Structure is RMSD able lowered by 7.7 Å, the value of Rg able lowered by 3.41 Å and value SASA able lowered by 1504 Å2. For the beta-sheet structure, is able to retain about 16% compared to wild-type who did not have the beta-sheet structure. For the alpha-helix structure, mutan-1 did not influence alpa-helix composition. The resulted mutant design will be used as a suggestion to engineer BsL mutant in wet experiment.