Chemical glycobiology. Part A, Synthesis, manipulation and applications of glycans

Başlık:

ISBN:

9780128114704

Yayın Bilgileri:

Cambridge : Academic Press, [2017]

Fiziksel Tanımlama:

1 online resource (xix, 409 pages) : illustrations (some color)

Series:

Methods in enzymology ; v. 597

Methods in enzymology ; v. 597.

Contents:

Front Cover; Chemical Glycobiology Part A. Synthesis, Manipulation and Applications of Glycans; Copyright; Contents; Contributors; Preface; References; Section I: Evolution and Engineering of Glycans and Glycan Processing Enzymes; Chapter One: Discovery of New Glycosidases From Metagenomic Libraries; 1. Introduction; 2. DNA Isolation and Purification; 2.1. Indirect DNA Isolation; 2.2. Direct DNA Isolation; 2.2.1. Equipment; 2.2.2. Buffers and Reagents; 2.2.3. Protocol; 2.3. DNA Purification; 3. Library Generation; 3.1. Large Insert Fosmid Libraries; 3.1.1. Equipment

3.1.2. Buffers and Reagents3.1.3. Protocol; 3.1.3.1. DNA End Repair; 3.1.3.2. Size Selection; 3.1.3.3. Ligation; 3.1.3.4. Phage Packaging and Transduction; 3.2. Small Insert Library; 3.2.1. Pipette Tip Shearing of DNA; 3.2.2. Enzymatic DNA Digestion; 3.2.3. Ligation and Transformation; 4. Library Screening; 4.1. Equipment; 4.2. Buffers and Reagents; 4.3. Protocol; 5. Preparation for Sequencing; 5.1. Fosmid Isolation; 5.1.1. Equipment; 5.1.2. Buffers and Reagents; 5.1.3. Protocol; 6. Concluding Remarks; Acknowledgments; References

Chapter Two: Structure-Guided Directed Evolution of Glycosidases: A Case Study in Engineering a Blood Group Antigen-Cleav ... 1. Introduction; 2. Structure-Guided Library Design and Generation; 2.1. Identification of Amino Acid Target Residues From a Glycosidase Structure; 2.2. Generating Glycosidase Mutant Libraries Through Site-Saturation Mutagenesis (by Overlap Extension); 2.2.1. Equipment; 2.2.2. Reagents and Buffers; 2.2.3. Procedure; 2.2.4. Notes; 2.3. Generating Glycosidase Mutant Libraries Through Error-Prone PCR; 2.3.1. Equipment; 2.3.2. Reagents and Buffers; 2.3.3. Procedure

2.3.4. Notes3. Enzymatic Preparation of Fluorogenic Oligosaccharides as Substrates for High-Throughput Assays; 3.1. In Vitro Enzymatic Synthesis of Fluorogenic Oligosaccharides; 3.1.1. Equipment; 3.1.2. Buffers and Solutions; 3.1.3. Procedure; 3.1.4. Notes; 3.2. Biosynthesis of Fluorogenic Oligosaccharides in Metabolically Engineered Bacteria; 4. Performing Enzymatic Assays; 4.1. High-Throughput Assays of Glycosidase Variants Expressed From Mutant Libraries; 4.1.1. Equipment; 4.1.2. Buffers, Solutions, and Media; 4.1.3. Procedure; 4.1.4. Notes

4.2. Determining the Enzyme Kinetics of Mutant Glycosidases4.2.1. Equipment and Software Tools; 4.2.2. Buffers, Solutions, and Reagents; 4.2.3. Procedure; 4.2.4. Notes; 5. Summary and Conclusion; Acknowledgments; References; Chapter Three: A Pipeline for Studying and Engineering Single-Subunit Oligosaccharyltransferases; 1. Introduction; 2. Materials; 2.1. Media; 2.2. Media Supplements; 2.3. Bacterial Strains and Plasmids; 2.4. GlycoSNAP Assay; 2.5. Preparation of ssOSTs by CFPS; 2.5.1. S30 Extract Preparation; 2.5.2. Producing ssOST in CFPS Supplemented With POPC Nanodiscs

Abstract:

Awareness of the biological significance of glycans and glycoconjugates continues to grow by leaps and bounds. Taking a cue from chemical biology, in general, the volumes in this two part treatment present strategies and methods from diverse fields of biochemistry, biophysics, bioinformatics, organic chemistry, and engineering, to examine glycans and their myriad functions. Volumes 597 and 598 encompass two interrelated areas: Synthesis, manipulation and applications of glycans B (v.597), and, Monitoring glycans and their interactions (v.598). The central theme of volume 597 concerns enzymes that assemble and process glycans and glycoconjugates. Glycans play a wide range of fundamental roles in all aspects of life in mono- and multicellular organisms, and the essentiality and diversity of carbohydrates in biology are no longer well-kept secrets. The development of these volumes on the theme of chemical glycobiology spotlights the significance and impact chemical biology approaches have on the advancement of glycobiology.

Local Note:

Elsevier

Subject Term:

SCIENCE -- Life Sciences -- Biochemistry.

Added Author: