Natural Product Biosynthesis by Microorganisms and Plants, Part A

1st Edition, Volume 515 - October 1, 2012
Latest edition
Editor: David A. Hopwood
Language: English

This new volume of Methods in Enzymology continues the legacy of this premier serial by containing quality chapters authored by leaders in the field. The first of 3 volumes coveri… Read more

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This new volume of Methods in Enzymology continues the legacy of this premier serial by containing quality chapters authored by leaders in the field. The first of 3 volumes covering Natural product biosynthesis by microorganisms and plants, it has chapters on such topics as Kinetics of plant sesquiterpene synthases, Terpenoid biosynthesis in fungi, and plant Type III polyketide synthases.

Series Page
Contributors
Preface
Methods in Enzymology
Chapter One Steady-State Kinetic Characterization of Sesquiterpene Synthases by Gas Chromatography–Mass Spectroscopy

1 Introduction
2 Experimental Components and Considerations
3 Pilot Experiments
4 Steady-State Kinetic Experiments
5 Data Handling/Processing
6 Summary

Chapter Two Automating Gene Library Synthesis by Structure-Based Combinatorial Protein Engineering

1 Overview
2 Plasmid Library Synthesis
3 Optimization of SCOPE PCR Conditions
4 Generic SCOPE Method
5 Applications of SCOPE Synthesis
6 Troubleshooting
7 Conclusions

Chapter Three In Planta Transient Expression Analysis of Monoterpene Synthases

1 Introduction
2 Plant Transformation Vector Construction
3 Agrobacterium-Mediated Transient Expression
4 Agrobacterium Preparation for High-Throughput Analysis
5 Product Identification
6 Transient Expression of a Kiwifruit Linalool Synthase
7 Conclusions

Chapter Four Natural Rubber Biosynthesis in Plants

1 Introduction
2 Preparing Samples for Assaying Rubber Transferase Activity
3 The Rubber Transferase Assay
4 Identification and Purification of Rubber Transferase
5 Qualitative Protein Analysis
6 Protein Quantification
7 Summary

Chapter Five Discovery and Characterization of Terpenoid Biosynthetic Pathways of Fungi

1 Introduction
2 Identification, Cloning, and Characterization of Terpene Synthases
3 Structural Characterization and Engineering of Fungal Terpene Synthase Activities
4 Identification and Characterization of Biosynthetic Gene Clusters
5 Genome Mining for Sesquiterpene Synthases
6 Conclusions

Chapter Six Menaquinone Biosyntheses in Microorganisms

1 Introduction
2 Futalosine Pathway
Chapter Seven Diversity and Analysis of Bacterial Terpene Synthases
1 Terpenoid Metabolites from Bacterial Cultures
2 Bacterial Terpene Synthases
3 Methods for the Study of Bacterial Terpene Synthases

Chapter Eight Platensimycin and Platencin Biosynthesis in Streptomyces platensis, Showcasing Discovery and Characterization of Novel Bacterial Diterpene Synthases

1 Introduction
2 Methods
3 Conclusions

Chapter Nine Strategies for Engineering Plant Natural Products

1 Introduction
2 Metabolic Engineering Strategies
3 Conclusions and Future Directions

Chapter Ten Discovery and Functional Analysis of Monoterpenoid Indole Alkaloid Pathways in Plants

1 Introduction
2 Enzyme Assays for MIA Pathway Steps
3 Methods for MIA Pathway Gene Discovery in C. roseus
4 Summary

Chapter Eleven Biochemical Genomics for Gene Discovery in Benzylisoquinoline Alkaloid Biosynthesis in Opium Poppy and Related Species

1 Benzylisoquinoline Alkaloids
2 Transcriptomics
3 Proteomics
4 Metabolomics
5 Integrated Functional Genomics

Chapter Twelve Analysis and Modification of Ergot Alkaloid Profiles in Fungi

1 Introduction
2 Extraction of Ergot Alkaloids from Biological Sources
3 Analysis of Ergot Alkaloids by HPLC
4 Identification of Candidate Genes via Comparative Genomics
5 Modification of Ergot Alkaloid Pathways via Transformation
6 Summary

Chapter Thirteen Engineering of Glucosinolate Biosynthesis

1 Introduction
2 Pathway Elucidation and Candidate Identification In Silico
3 Metabolic Engineering of Glucosinolates in N. benthamiana
4 Metabolic Engineering of Glucosinolates in S. cerevisiae
5 Conclusion

Chapter Fourteen Structure–Function Analyses of Plant Type III Polyketide Synthases

1 Introduction
2 Bioinformatic Analyses of Type III PKSs
3 In Vivo Biochemical Function of Type III PKSs
4 Expression of Type III PKSs
5 Structural Analyses of Type III PKSs
6 Kinetic Analyses of Type III PKSs
7 Mutagenic Exploration of Type III PKS Function and Adaptive Evolution

Chapter Fifteen Engineering of Plant Type III Polyketide Synthases

1 Introduction
2 Benzalacetone Synthase
3 Pentaketide Chromon Synthase
4 Octaketide Synthase
5 Polyketide Synthase 1
6 Practical Considerations
7 Summary and Future Prospects

Chapter Sixteen Type III Polyketide Synthases in Microorganisms

1 Introduction
2 Methods of Study

Author Index
Subject Index

Life Sciences

Physical Sciences & Engineering

Social Sciences & Humanities

Health

Natural Product Biosynthesis by Microorganisms and Plants, Part A

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David A. Hopwood

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