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Chapter 7 : Strategies for Accessing Microbial Secondary Metabolites from Silent Biosynthetic Pathways
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This chapter concentrates on the methodological approaches used to access the products of silent biosynthetic pathways (SBPs), and presents a number of important case studies that are intended to help one evaluate the advantages and disadvantages of each technique. Usage of the term ‘’silent’’ throughout this chapter is intended to include the broad range of suppressive transcriptional, translational, and biosynthetic mechanisms responsible for inhibiting microbial secondary-metabolite formation. During the preparation of this chapter, the authors have found it extremely useful to consider each method in its chronological context. An exciting new dimension in SBP studies is the emergence of novel mechanism-based and other hybrid techniques that meld the technological simplicity of culture manipulation strategies with recent breakthroughs in the understanding of microbial molecular biology. Although significant insight from genome mining was used to advance this discovery, this work serves as a testament to the fact that, in the absence of similar guiding factors, culture modification strategies will continue to involve significant guesswork regarding which culture amendments will provide access to SBPs. Genomic technologies are expected to facilitate the sequencing of microbial genomes at even greater rates, which is anticipated to provide natural products researchers with unprecedented capacity for identifying SBPs. Currently available techniques for investigating SBPs fall into three discrete categories: (i) culture-dependent, (ii) genomics- driven, and (iii) mechanism-based and hybrid approaches. Each of these method sets present unique advantages and disadvantages for studying SBPs.
Key Concept Ranking
- High-Performance Liquid Chromatography
Timeline illustrating significant events related to the field of SBP studies. Note the substantial surge in the number of experimental methods and techniques for investigating SBPs that were developed during the past several years.
Representative microbial compounds identified by use of early culture manipulation techniques.
Compounds obtained from Sphaeropsidales sp. F-24′707 by use of culture manipulation.
Compounds obtained from A. ochraceus DSM7428 by use of culture manipulation.
Examples of natural products obtained by manipulating microbial culture conditions.
Examples demonstrating the use of coculture techniques to induce the production of microbial natural products.
Combined approach leading to the discovery of coelichelin from S. coelicolor.
Examples of microbial metabolites whose structures were predicted based on genome scanning. The structure of each metabolite was later confirmed by isolation and characterization using NMR and mass spectrometric analyses.
Examples of SBP-derived natural products that were identified by use of heterologous expression techniques.
Compounds obtained from SBPs following activation using promoter/activator manipulation.
Natural products obtained by using the chemical epigenetic induction of SBPs.
Overview of the genomisotopic method and its application to the study of the orfamide gene cluster in P. fluorescens Pf-5.
Numbers of PKS, NRPS, and HPN gene clusters in selected bacteria and fungi
Examples of some reported culture manipulation techniques
Examples of cocultivation methods for investigating SBPs in microorganisms
Examples of online resources providing archiving of microbial genomic data a
Summary of the major strengths and weaknesses of methods for investigating microbial SBPs