Natural Product Biosynthesis by Microorganisms and Plants Part B

Chapter One. In Vivo Production of Thiopeptide Variants

1 Introduction

2 Generation of Thiopeptide Variants

References

Chapter Two. Microviridin Biosynthesis

1 Introduction

2 Reconstitution In Vitro of the Cyclization Reactions of the Microviridin Biosynthetic Pathway from P. agardhii

3 Influence of Leader Peptide on the Cyclization Reactions

4 Summary

References

Chapter Three. Cyclotide Isolation and Characterization

1 Introduction

2 Isolation of Cyclotides from Plant Tissues

3 Isolation of Nucleic Acids Encoding Cyclotide Precursors

4 Mass Spectrometric Detection and Characterization of Cyclotides

5 Structural Analysis of Cyclotides

6 Membranolytic Assays of Cyclotides

7 Summary

References

Chapter Four. Ribosomally Encoded Cyclic Peptide Toxins from Mushrooms

1 Introduction

2 Detection and Purification of Amanita Cyclic Peptide Toxins

3 Identification of Cyclic Peptide Genes in Amanita and Other Fungi

4 Purification and Assay of Prolyl Oligopeptidase (POP) from Mushrooms

5 Immunodetection of POP in Mushroom Tissues

6 Summary

References

Chapter Five. The Pictet–Spengler Mechanism Involved in the Biosynthesis of Tetrahydroisoquinoline Antitumor Antibiotics

1 Introduction

2 Biosynthetic Analysis of THIQ Core Scaffold Assembly

3 Reaction Mechanism for Constructing a Pentacyclic THIQ Core Skeleton Catalyzed by a Novel Function for a Single NRPS Module

4 Comparison Between Typical Peptide Synthesis and the Pictet–Spengler Reaction Catalyzed by NRPS Machinery

5 Reaction Profiles Affected by the Chain Length of the Acyl Moiety

6 Multiple Roles for the Fatty Acyl Chain During the Maturation Process in Late-Stage Biosynthesis

7 Methods for Substrate Synthesis, Enzyme Preparation, and Assay Conditions

8 Conclusion

References

Chapter Six. Discovery and Biosynthesis of Phosphonate and Phosphinate Natural Products

1 Introduction

2 Spectrum of Phosphonate Natural Products: Structures and Bioactivities

3 Biosynthetic Pathways

4 Screening Bacteria for Phosphonate Producing Gene Clusters

5 Purification of Phosphonates/Phosphinates

6 Structural Elucidation of Phosphonates

7 A Wealth of Unusual Biosynthetic Enzymes

8 Conclusion and Outlook

References

Chapter Seven. RlmN and AtsB as Models for the Overproduction and Characterization of Radical SAM Proteins

1 Introduction

2 RlmN as a Model for Gene Expression and Overproduction of RS Proteins

3 RlmN as a Model for Purification of RS Proteins

4 Overproduction and Purification of E. coli Flavodoxin and Flavodoxin Reductase

5 AtsB as a Model for Determination of Configuration and Stoichiometry of Iron–Sulfur Clusters in RS Enzymes

6 Activity Determination of RS Enzymes

References

Chapter Eight. Fe(II)-Dependent, Uridine-5′-Monophosphate α-Ketoglutarate Dioxygenases in the Synthesis of 5′-Modified Nucleosides

1 Introduction

2 Methods

3 Summary

References

Chapter Nine. Tailoring Reactions Catalyzed by Heme-Dependent Enzymes

1 Introduction

2 Overexpression, Purification, and Characterization of the Novel Nitrating CYP TxtE

3 Protein Characterization Using UV–vis Spectroscopy

4 Ligand and Substrate Binding

5 Summary

References

Chapter Ten. Oxidative Tailoring Reactions Catalyzed by Nonheme Iron-Dependent Enzymes

1 Introduction

2 Examples of Tailoring Reactions Catalyzed by Nonheme Iron- and Oxygen-Dependent Enzymes

3 Heterologous Expression and Functional Investigation of the S. coelicolor redH and redG Genes in Streptomyces albus

4 Conclusions

References

Chapter Eleven. The Rare Fluorinated Natural Products and Biotechnological Prospects for Fluorine Enzymology

1 Introduction

2 Production of Fluoroacetate and 4-Fluorothreonine from S. cattleya

3 Enzyme 1: Fluorinase

4 Enzyme 2: Purine Nucleoside Phosphorylase

5 Enzyme 3: Isomerase

6 Enzyme 4: Aldolase

7 Enzyme 5: Transaldolase (Pyridoxal Phosphate)

8 Enzyme 6: Aldehyde Dehydrogenase

9 Generation of a Novel Fluorometabolite by Genetic Engineering

10 Summary

References

Chapter Twelve. Enzymatic Chlorination and Bromination

1 Introduction

2 FADH2-Dependent Halogenases

3 Characterization of FADH2-Dependent Halogenases

4 Nonheme Iron, α-Ketoglutarate- and O2-Dependent Halogenases

References

Chapter Thirteen. Prenyltransferases of the Dimethylallyltryptophan Synthase Superfamily

1 Introduction

2 Spore Preparation

3 Cultivation of Fungi

4 RNA and DNA Isolation and cDNA Synthesis

5 Gene Cloning

6 Protein Overproduction

7 Preparation of Stock Solutions for Enzyme Assays

8 Biochemical Characterization of Prenyltransferases

9 Chemoenzymatic Synthesis of Prenylated Compounds

10 HPLC

11 Summary

References

Chapter Fourteen. Serine Carboxypeptidase-Like Acyltransferases from Plants

1 Introduction

2 Methods in SCPL Acyltransferase Research

3 Summary

References

Chapter Fifteen. 4-Methylideneimidazole-5-One-Containing Aminomutases in Enediyne Biosynthesis

1 Introduction

2 Methods

3 Conclusion

References

Chapter Sixteen. Tailoring Enzymes Acting on Carrier Protein-Tethered Substrates in Natural Product Biosynthesis

1 Introduction

2 Methods

3 Conclusion

References

Chapter Seventeen. High-Throughput Colorimetric Assays for Nucleotide Sugar Formation and Glycosyl Transfer

1 Introduction

2 Purification of the Glycosyltransferase OleD

3 Colorimetric Assay for NDP-Sugar Formation

4 Coupled Reactions for Glycosyl Transfer

5 High-Throughput Colorimetric Coupled Assay for Glycosyl Transfer

6 Summary

References

Author Index

Subject Index