Format:
1 Online-Ressource (538 Seiten)
Edition:
First edition
ISBN:
9780128105030
Series Statement:
Methods in enzymology volume 590
Content:
Nanoarmoring of Enzymes: Rational Design of Polymer-Wrapped Enzymesis the latest volume in the Methods in Enzymologyseries and focuses on nanoarmoring of enzymes and the rational design of polymer-wrapped enzymes.Focuses on the nanoarmoring of enzymesCovers the rational design of polymer-wrapped enzymesIncludes contributions from leading authorities working in enzymologyInforms and updates on all the latest developments in the field of enzymology
Content:
Front Cover -- NanoArmoring of Enzymes: Rational Design of Polymer-Wrapped Enzymes -- Copyright -- Contents -- Contributors -- Preface -- References -- Chapter One: Encapsulating Proteins in Nanoparticles: Batch by Batch or One by One -- 1. Introduction -- 2. "Batch-by-Batch" Encapsulation of Proteins in Silica NPs -- 2.1. A Facile General Noncovalent Method to Armor Proteins and Peptides in Silica NPs -- 2.1.1. Protocol -- 2.1.1.1. Chemicals and Materials -- 2.1.1.1.1. Equipment -- 2.1.1.2. Procedures -- 2.1.1.2.1. Tips -- 2.1.2. Application Examples: Armord EGFP as a Robust Fluorescent Probe -- 2.2. An Efficient Covalent Method to Armor Proteins in Silica NPs -- 2.2.1. Protocol -- 2.2.1.1. Chemicals and Materials -- 2.2.1.1.1. Equipment -- 2.2.1.2. Procedures -- 2.2.1.2.1. Tips -- 2.2.2. Application Examples -- 2.2.2.1. Case I: In Situ Synthesis of Porous Silica NPs for Immobilization of Catalase -- 2.2.2.2. Case II: Encapsulated EGFP for Cellular Imaging -- 3. "One-by-One" Encapsulation of Proteins in NPs -- 3.1. Individually Coating Protein Molecules in Silica NPs -- 3.1.1. Protocol -- 3.1.1.1. Chemicals and Materials -- 3.1.1.1.1. Equipment -- 3.1.1.2. Procedures -- 3.1.1.2.1. Tips -- 3.1.2. Application Examples -- 3.1.2.1. Individually Coating NIRFP in Silica NPs for Bioimaging -- 3.2. Individually Coating Protein Molecules in CS NPs -- 3.2.1. Protocol -- 3.2.1.1. Materials and Chemicals -- 3.2.1.1.1. Equipment -- 3.2.1.2. Procedures -- 3.2.1.2.1. Tips -- 3.2.2. Application Example -- 3.2.2.1. CS-Coated Red Fluorescent Protein as a Dual-Functional siRNA Carrier -- 4. Summary and Outlook -- Acknowledgments -- References -- Chapter Two: Enzyme Adsorption on Nanoparticle Surface Probed by Highly Sensitive Second Harmonic Light Scattering -- 1. Introduction -- 2. Techniques for Probing Nanoparticle-Protein Interaction
Content:
3. Second Harmonic Light Scattering -- 3.1. Background -- 3.2. SHLS From GNPs -- 3.3. Apparatus -- 4. Sample Preparation -- 4.1. Gold Nanoparticles -- 4.2. Proteins -- 5. Interaction of ADH With GNPs -- 6. Interaction of Insulin With GNPs -- 7. Stoichiometry of the Protein-GNP Conjugate -- 8. Summary and Future Outlook -- Acknowledgment -- References -- Chapter Three: Armoring Enzymes by Metal-Organic Frameworks by the Coprecipitation Method -- 1. Introduction -- 2. Equipment -- 3. Materials -- 4. Protocol -- 4.1. Duration -- 4.2. Preparation -- 5. Safety -- 6. Step1: Preparation of EnzymeMOF -- 6.1. Overview -- 6.2. Methods -- 6.3. Tip -- 6.4. Tip -- 6.5. Tip -- 7. Step2: Preparation of Buffers -- 7.1. Overview -- 7.2. Methods -- 7.3. Tip -- 7.4. Tip -- 8. Step3: Purification of the Product -- 8.1. Overview -- 8.2. Methods -- 8.3. Tip -- 8.4. Tip -- 9. Step4: Characterization of Enzyme@MOF -- 9.1. Overview -- 9.2. Methods -- 9.3. Tip -- 9.4. Tip -- 10. Step5: Measurement of Enzyme Loading in MOFS -- 10.1. Overview -- 10.2. Methods -- 10.3. Tip -- 10.4. Tip -- 10.5. Tip -- 11. Step6: Analysis of the Location of Enzymes in MOFS -- 11.1. Overview -- 11.2. Methods -- 11.3. Tip -- 11.4. Tip -- 11.5. Tip -- 12. Step7: Determination of Enzyme Activities -- 12.1. Overview -- 12.2. Methods -- 12.3. Tip -- 13. Conclusions -- References -- Chapter Four: Enzyme Armoring by an Organosilica Layer: Synthesis and Characterization of Hybrid Organic/Inorganic Nanobi ... -- 1. Introduction -- 2. Equipments and Chemicals -- 3. Safety Guidelines -- 4. Procedures -- 4.1. General Synthetic Strategy at a Glance -- 4.2. Silica Nanoparticles Synthesis -- 4.2.1. Silica Nanoparticle Synthesis Protocol -- 4.2.2. Silica Nanoparticles Characterization -- 4.3. Tips on Silica Nanoparticle Synthesis -- 4.4. Enzyme Immobilization and Shielding
Content:
4.4.1. General Procedure and Experimental Set-Up -- 4.4.2. Silica Nanoparticle Functionalization -- 4.4.3. Enzyme Crosslinking and Shielding -- 4.4.4. Nanobiocatalyst Characterization -- 4.4.4.1. Temperature Stress -- 4.4.4.2. Ultrasound Stress -- 4.4.4.3. pH Stress -- 4.4.4.4. Chaotropic Stresses -- 4.4.4.5. Proteinase K Digestion -- 4.4.5. Shielding of Different Enzymes -- 4.5. Tips on the Immobilization Yield and on the Activity of the Immobilized Enzyme -- 4.6. Tips on the Curing of the Shielded Enzymes -- 4.7. Tips on the Calculation of the Carrier Surface -- 4.8. Tips for a Successful Enzyme Crosslinking and Shielding -- 4.9. Tips on Particle Loss -- 5. Conclusions -- Acknowledgments -- References -- Chapter Five: Strategies for Biophysical Characterization of Protein-Polymer Conjugates -- 1. Introduction -- 2. Biophysical Methods -- 2.1. Differential Scanning Fluorimetry -- 2.1.1. Summary -- 2.1.2. Advantages and Disadvantages -- 2.1.3. Sample Requirements and Preparation -- 2.1.4. Instrumentation and Consumables -- 2.1.4.1. Thermocycler -- 2.1.4.2. Plates -- 2.1.4.3. Solvatofluorochromic Dye -- 2.1.5. Protocol -- 2.1.6. Data Analysis and Typical Data -- 2.2. Intrinsic Tryptophan Fluorescence -- 2.2.1. Summary -- 2.2.2. Advantages and Disadvantages -- 2.2.3. Sample Requirements and Preparation -- 2.2.4. Instrumentation and Consumables -- 2.2.4.1. Fluorimeter -- 2.2.4.2. Plates -- 2.2.4.3. Buffers and Denaturant -- 2.2.5. Protocol -- 2.2.6. Data Analysis and Typical Data -- 3. Conclusions -- Acknowledgments -- References -- Chapter Six: Guide to the Preparation of Molecularly Imprinted Polymer Nanoparticles for Protein Recognition by Solid-Pha ... -- 1. Introduction -- 2. Protocols -- 2.1. Activation of GBs -- 2.2. Functionalization of GBs -- 2.3. Enzyme Immobilization and Synthesis of MIP-NPs -- 3. Physicochemical Characterization of MIP-NPs
Content:
4. Evaluation of the Binding Characteristics of MIP-NPs Using a Quartz Crystal Microbalance -- 5. MIP-NPs Protect Enzyme From Thermal and pH Denaturation -- 6. Concluding Remarks -- 7. Notes -- Acknowledgments -- References -- Chapter Seven: Armored Urease: Enzyme-Bioconjugated Poly(acrylamide) Hydrogel as a Storage and Sensing Platform -- 1. Introduction -- 2. Reversible Immobilization -- 3. Jack Bean Urease: Active Site -- 4. Immobilization of Jack Bean Urease -- 5. Methodology -- 5.1. Equipment and Reagents -- 5.1.1. General Equipment -- 5.1.2. Preparation of Hydrogel -- 5.1.3. Immobilization of Urease in the Hydrogel -- 5.1.4. Urease-Urea Assay -- 5.1.5. Characterization Instrumentation -- 5.1.6. Preparation of Crystalline Colloidal Array -- 5.2. Preparation of Hydrogel -- 5.3. Immobilization of Urease in the Hydrogel -- 5.4. Urease-Urea Assay -- 5.5. Analysis of Urease-Urea Assay With UCG and Free Urease -- 5.5.1. Michaelis-Menten Constant (Km) Values -- 5.5.2. Effect of pH, Thermal History, and Thermal Inactivation Studies -- 5.6. Characterization Methods -- 5.7. Methodology for Sensing Mercury -- 5.7.1. Preparation of Crystalline Colloidal Array -- 5.7.2. Preparation of Polymerized Crystalline Colloidal Array -- 5.7.3. Preparation of Ion Strength Responsive PCCA (Carboxylated PCCA) -- 5.7.4. Preparation of Urease-Coupled PCCA Sensor for Mercury -- 5.7.5. Measurement of UPCCA Mercury Sensor Response -- 6. Results and Discussion -- 6.1. Urease Stability at Room Temperature: Comparison Between Free Urease and Immobilized Urease -- 6.2. Effect of pH and Temperature on the Enzyme-Coupled Hydrogel Matrix (UCG) -- 6.3. Sensing of Mercury Pollutant in Water -- 7. Conclusions and Future Perspectives -- Acknowledgments -- References -- Chapter Eight: Armored Enzyme-Nanohybrids and Their Catalytic Function Under Challenging Conditions -- 1. Introduction
Content:
1.1. Need for Enzyme Armor -- 1.2. Synthesis of Bienzyme-Polymer Conjugates -- 1.3. Adsorption of the Bienzyme-Polymer Onto GO -- 2. Methods: Synthesis of Bienzyme-Polymer Conjugates -- 2.1. Materials -- 2.1.1. Equipment -- 2.1.2. Reagents -- 2.2. Synthesis of Bienzyme-Polymer Conjugates -- 3. Methods: Synthesis of Bienzyme-Polymer-Graphene Oxide (GOx-HRP-PAA/GO) Hybrid Materials -- 3.1. Materials -- 3.1.1. Equipment -- 3.1.2. Reagents -- 3.2. Synthesis of Bienzyme-Polymer GO Hybrid Materials -- 4. Characterization -- 4.1. Agarose Gel Electrophoresis -- 4.1.1. Equipment -- 4.1.2. Reagents -- 4.2. Zeta Potential -- 4.2.1. Equipment -- 4.2.2. Materials -- 4.2.3. Procedure -- 4.3. Transmission Electron Microscopy -- 4.3.1. Equipment -- 4.3.2. Materials and Reagents -- 4.4. Circular Dichroism Spectroscopy -- 4.4.1. Equipment -- 4.4.2. Materials -- 4.4.3. Procedure -- 4.4.4. Results -- 4.5. Activity Studies -- 4.5.1. Equipment -- 4.5.2. Materials -- 4.5.3. Activity Studies at 25°C at Multiple pH Values -- 4.5.4. Activity Studies at 65°C and in the Presence of a Denaturant (4 mM SDS) -- 5. Conclusions -- Acknowledgments -- References -- Chapter Nine: Approaches for Conjugating Tailor-Made Polymers to Proteins -- 1. Introduction -- 1.1. Background Information -- 1.2. General Considerations in Polymer Synthesis, Conjugation, and Characterization -- 1.2.1. Safety Considerations -- 1.2.2. Personal Protective Equipment -- 1.2.3. Materials Needed -- 1.2.4. Equipment Needed -- 2. Polymer Synthetic Procedures -- 2.1. RAFT CTAs and ATRP Initiators -- 2.1.1. Synthesis of PAETC -- 2.1.2. Synthesis of PYPETC -- 2.2. Typical Polymerization Methods -- 2.2.1. Typical RAFT Polymerization of Acrylamide Using PAETC -- 2.2.2. Precipitation of Am Polymers -- 2.2.3. Typical RAFT Polymerization of N,N-Dimethylacrylamide Using PAETC -- 2.2.4. Precipitation of DMAm Polymers
Content:
2.2.5. RAFT Polymerization With PYPETC
Note:
Inclou referències bibliogràfiques i índex
Additional Edition:
ISBN 9780128105023
Additional Edition:
Erscheint auch als Druck-Ausgabe Kumar, Challa Vijaya NanoArmoring of Enzymes: Rational Design of Polymer-Wrapped Enzymes Saint Louis : Elsevier Science,c2017 ISBN 9780128105023
Language:
English
Subjects:
Biology
Keywords:
Enzym
;
Design
;
Nanopartikel
Bookmarklink
