Nonribosomal Peptide and Polyketide Biosynthesis: Methods and Protocols

Description: This volume presents an overview of nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) structure and function. It then continues with methods for the analysis of these pathways including conventional enzymological assays, contemporary mass spectrometric analysis techniques, specialized molecular biological approaches applicable to NRPSs and PKSs, and small molecule analysis tools tailored to this very special class of natural products, and concludes by examining bioinformatics tools for the analysis of these enzymes, pathways, and molecules. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Nonribosomal Peptide and Polyketide Biosynthesis: Methods and Protocols serves as a valuable reference for those experienced in studying NRPS and PKS enzymes, pathways, and natural products as well as a gateway for those just entering the field. Part I: Background and Overview 1. Structural Biology of Nonribosomal Peptide Synthetases Bradley R. Miller and Andrew M. Gulick 2. The Assembly Line Enzymology of Polyketide Biosynthesis Marisa Till and Paul R. Race Part II: In Vitro Methods 3. Measurement of Nonribosomal Peptide Synthetase Adenylation Domain Activity Using a Continuous Hydroxylamine Release Assay Benjamin P. Duckworth, Daniel J. Wilson, and Courtney C. Aldrich 4. Affinity Purification Method for the Identification of Nonribosomal Peptide Biosynthetic Enzymes Using a Synthetic Probe for Adenylation Domains Fumihiro Ishikawa and Hideaki Kakeya 5. Colorimetric Detection of the Adenylation Activity in Nonribosomal Peptide Synthetases Chitose Maruyama, Haruka Niikura, Masahiro Takakuwa, Hajime Katano, and Yoshimitsu Hamano 6. Facile Synthetic Access to Glycopeptide Antibiotic Precursor Peptides for the Investigation of Cytochrome P450 Action in Glycopeptide Antibiotic Biosynthesis Clara Brieke, Veronika Kratzig, Madeleine Peschke, and Max J. Cryle 7. Reconstitution of Fungal Nonribosomal Peptide Synthetases in Yeast and In Vitro Ralph A. Cacho and Yi Tang 8. The Continuing Development of E. coli as a Heterologous Host for Complex Natural Product Biosynthesis Haoran Zhang, Lei Fang, Marcia S. Osburne, and Blaine A. Pfeifer 9. Screening for Expressed Nonribosomal Peptide Synthetases and Polyketide Synthases Using LC¿MS/MS-Based Proteomics Yunqiu Chen, Ryan A. McClure, and Neil L. Kelleher 10. Enhancing Nonribosomal Peptide Biosynthesis in Filamentous Fungi Alexandra A. Soukup, Nancy P. Keller, and Philipp Wiemann 11. In Situ Analysis of Bacterial Lipopeptide Antibiotics by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging Delphine Debois, Marc Ongena, Hélène Cawoy, and Edwin De Pauw 12. Secondary Metabolic Pathway-Targeted Metabolomics Maria I. Vizcaino and Jason M. Crawford Part III: Bioinformatic Methods 13. Annotating and Interpreting Linear and Cyclic Peptide Tandem Mass Spectra 14. Bioinformatics Tools for the Discovery of New Nonribosomal Peptides Valérie Leclère, Tilmann Weber, Philippe Jacques, and Maude Pupin 15. The Use of ClusterMine360 for the Analysis of Polyketide and Nonribosomal Peptide Biosynthetic Pathways Nicolas Tremblay, Patrick Hill, Kyle R. Conway, and Christopher N. Boddy 16. Alignment-Free Methods for the Detection and Specificity Prediction of Adenylation Domains Guillermin Agüero-Chapin, Gisselle Pérez-Machado, Aminael Sánchez-Rodríguez, Miguel Machado Santos, and Agostinho Antunes 17. Characterization of Nonribosomal Peptide Synthetases with NRPSsp Carlos Prieto

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