THE EFFECT VARIATION OF SOLVENT ON LIPASE TRANSESTERIFICATION ACTIVITY: EXPERIMENTAL AND BIOCOMPUTIONAL APPROACHES

• Main Author: Anzar Hamzah, Tiandi
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/64732
• Institution: Institut Teknologi Bandung

The transesterification reaction of lipase occurs when an organic solvent is used. The study aimed to determine the transesterification activity of local isolates of lipases and commercial lipases using experimental and biocomputational approaches in organic solvents. The enzymes used consisted of local isolate lipase LK_ITB5a and commercial lipase Lipozyme TL IM. The transesterification activity of lipase was tested by synthesizing pNP esters using methyl palmitate as a substrate in acetone, acetonitrile (ACN), and n-hexane as a solvent. The transesterification assay was carried out at 120 rpm shaking, 50 °C temperature, and time intervals of 2, 4, and 6 hours. Using a UV-VIS spectrophotometer set to 410 nm, the reaction was examined for changes in pNP substrate concentration. The highest transesterification activity of the enzyme LK ITB5a at a total concentration of 0.50 mg/mL was 58.26.U.g-1 in acetone solvent for a reaction time of 6 hours, which was 71.92 times greater than the highest transesterification activity of Lipozyme TL IM at a total concentration of 1.49 mg/mL at 0.81.U.g-1 in acetonitrile solvent for a reaction time of 4 hours.Using the AutodockVina application, the protein-ligand docking process was carried out to examine the interaction between the receptor and the ligand during a catalytic reaction. GROMACS was used in a variety of solvents to optimize the geometry of the protein and ligand structures. The final stage is rescoring with the DCDFTB method to improve docking value accuracy using quantum mechanics theory. The results of the experiments revealed that LK_ ITB5a had the highest transesterification activity in acetone, followed by acetonitrile and n-hexane. In acetone, the enzyme LK_ITB5a has a hydrogen interaction between the ester oxygen atom on the substrate and the hydrogen atom in the imidazole group of residue H277, as well as a hydrogen interaction between the carbonyl oxygen of the substrate and the carbonyl oxygen atom on the S109 residue, where the interactions occur in accordance with the transesterification reaction mechanism. In acetonitrile, the LK_ITB5a enzyme has only one protein-ligand interaction, which corresponds to the transesterification reaction mechanism, whereas there is no protein-ligand interaction in n-hexane. The transesterification activity of the LK_ITB5a enzyme was confirmed computationally as a result of these findings. Meanwhile, the observed protein-ligand interactions in the Lipozyme TL IM enzyme in all solvent variations did not match the mechanism of the transesterification reaction. This indicates that the enzyme Lipozyme TL IM transesterification activity has not yet been computationally confirmed.