IN SILICO STUDY ON THERMOSTABILITY AND ACTIVITY IMPROVEMENTS OF PETASE AND MHETASE FROM IDEONELLA SAKAIENSIS

• Main Author: Zahroh Janatunaim, Rifqi
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/54200
• Institution: Institut Teknologi Bandung

Polyethylene terephthalate (PET) is a synthetic polyester which is widely produced due to its durability and convenient shape. Consequently, there was increasing in waste of PET which causes problems in the environment and human health because it hard to decompose. Degradation method using biological technique is more potential than chemical and mechanical degradation due to the environmentally friendly. Studies on biodegradation using enzymes have been carried out, one of most specific PET-degrading enzymes were PETase and MHETase derived from Gram-negative bacteria, Ideonella sakaiensis. Nevertheless, both enzymes are heat-labile and would lose the activity at temperatures above 40 ° C. Meanwhile, to enhance the effectiveness of the PET degradation require around 50–65 ° C temperature condition. Accordingly, study on thermostability and activity enhancement are required. Genetic engineering studies using wet lab method widely used to increase the effectiveness of enzymes. However, this method is less time and cost effective. In silico studies can be an additional method for effectiveness of wet lab by design, predict yields, and analyze the amino acid character prior to wet lab testing. Based on these, this study will design the combination of mutant using in-silico to obtain not only stable PETase and MHETase but also strong affinity and interaction with various ligands. In this study, PETase and MHETase mutant data were obtained by literature studies, mutation processes and stability analyzes were performed using FoldX software and molecular docking of enzyme-ligan would run using AutoDock software. The stability and molecular docking analysis revealed that PETaseL117F/Q119Y/S121E/G165A/D186H/R280A/S214H /S238F variant had -7.38 kcal/mol of ??G. This variant strongly interacts with the main substrat were PET and other polyesters were PEF and PBT. This study also demonstrated that MHETaseD304A/D307A/S267A/S419/F424N variant had -7.9 kcal/mol of ??G. This variant strongly interacts with the main substrats were MHET, BHET, and 2-HE(MHET)2 and low affinity interacting to other polyesters were PBT. The combined mutant design of PETase and MHETase which were stable and had strong affinity and interaction to various ligands can be obtained using in silico method. Thus, this study demonstrated the potential of in silico method to be developed in predicting research results and protein engineering designs to improve the quality of enzymes before working on the wet lab work.