In Silico Engineering of Disulphide Bonds to Produce Stable Cellulase (SpringerBriefs in Applied Sciences and Technology)

This Brief highlights different approaches used to create stable cellulase and its use in different fields. Cellulase is an industrial enzyme with a broad range of significant applications in biofuel production and cellulosic waste management. Cellulase 7a from Trichoderma reesei is the most efficient enzyme in the bio hydrolysis of cellulose. In order to improve its thermal stability, it can be engineered using a variety of approaches, such as hydrophobic interactions, aromatic interactions, hydrogen bonds, ion pairs and disulfide bridge creation. Preface 6 Contents 7 1 Introduction of Cellulase and Its Application 9 Abstract 9 1.1 Biofuel as a Suitable Alternative for Fossil Fuel 9 1.2 Enzyme Stability Is One of the Limits in Biofuel Production 11 1.3 A Procedure to in Silico Engineering of Cellulase 11 1.4 Cellulase 7a Selected as a Target 11 1.5 Advantages of Cellulose in Silico Engineering 12 References 12 2 Literature Review of Cellulase and Approaches to Increase Its Stability 14 Abstract 14 2.1 Cellulose and Cellulases Structure 14 2.2 Applications of Cellulase 15 2.2.1 Application of Cellulase in Biofuel Production 15 2.3 Trichoderma reesei Known as the Crowned King of Cellulolytic Fungi 17 2.4 Catalytic Mechanisms of Glycoside Hydrolases 17 2.5 An Introduction to Cellulase Family 7 19 2.6 Cellulose Biodegradation Procedures 20 2.7 Protein Engineering as a Solution 20 2.7.1 Rational Design Approach 21 2.8 Advantages of Enzyme Thermo Stabilization 23 2.9 Some Thermo Stabilize Mutations 23 2.9.1 Introducing Disulfide Bonds as a Strategy to Increase Stability 23 2.9.1.1 Mechanism of Disulfide Bond Formation 24 2.10 Molecular Dynamic Simulation as a Strong Approach to Evaluating Structure Stability 25 References 26 3 Methodology of Mutant Creation and Molecular Dynamic Simulation 29 Abstract 29 3.1 Obtaining Protein Structure File 29 3.2 Fixing pdb File 30 3.2.1 Removing Water Molecule and Ligands 30 3.3 Generation of Mutated pdb Files 30 3.4 Molecular Dynamic Simulation Steps 30 3.4.1 pdb2gmx (Topology File Generation) Tool 32 3.4.2 Unit Cell Defining the by the Tools, editconf and Adding Solvent 32 3.4.3 Addition of Ions 33 3.4.4 Energy Minimization as a Critical Step 33 3.4.5 Equilibration of Temperature and Pressure 34 3.4.6 MD Production 34 3.5 Tools for Analysis of Molecular Dynamics Trajectory 35 References 35 4 Result and Discussion of Molecular Dynamic Simulation of Created Mutants 36 Abstract 36 4.1 Nominated Residues for Mutation 36 4.2 Native Protein Structure and Stability Analysis 37 4.3 Mutated Protein Number 1 (E385C and A392C) Structure and Stability Analysis 38 4.4 Mutated Protein Number 2 (Y321C and A333C) Structure and Stability Analysis 42 4.5 Mutated Protein Number 3 (T383C and T399C) Structure and Stability Analysis 45 4.6 Mutated Protein Number 4 (A187C) Structure and Stability Analysis 46 4.7 Mutated Protein Number 5 (D257C and L346C) Structure and Stability Analysis 49 5 Conclusion of Simulation Analysis of Mutants 52 Abstract 52 5.1 Conclusion of MD Simulation of Created Mutants 52 5.2 Future Work of the Disulfide Bond Engineering to Cellulase 53 Acknowledgements 53 Front Matter....Pages i-viii Introduction of Cellulase and Its Application....Pages 1-5 Literature Review of Cellulase and Approaches to Increase Its Stability....Pages 7-21 Methodology of Mutant Creation and Molecular Dynamic Simulation....Pages 23-29 Result and Discussion of Molecular Dynamic Simulation of Created Mutants....Pages 31-46 Conclusion of Simulation Analysis of Mutants....Pages 47-48