Methods in enzymology. Volume459, Complex enzymes in microbial natural product biosynthesis. Part B, Polyketides, aminocoumarins and carbohydrates / edited by David A. Hopwood.

Format:

Book

Contributor:

Hopwood, D. A.

Series:

Methods in enzymology ; v. 459B.

Methods in enzymology, 0076-6879 ; v. 459B

Language:

English

Subjects (All):

Biosynthesis.

Polyketides--Synthesis.

Polyketides.

Peptides--Synthesis.

Peptides.

Physical Description:

1 online resource (644 p.)

Edition:

1st ed.

Place of Publication:

Amsterdam [Netherlands] ; Boston [Mass.] : Academic Press/Elsevier, 2009.

Language Note:

English

Summary:

Microbial natural products have been an important traditional source of valuable antibiotics and other drugs but interest in them waned in the 1990s when big pharma decided that their discovery was no longer cost-effective and concentrated instead on synthetic chemistry as a source of novel compounds, often with disappointing results. Moreover understanding the biosynthesis of complex natural products was frustratingly difficult. With the development of molecular genetic methods to isolate and manipulate the complex microbial enzymes that make natural products, unexpected chemistry has been

Contents:

Front Cover; Methods in Enzymology Complex Enzymes inMicrobial Natural ProductBiosynthesis, Part B:Polyketides,Aminocoumarins andCarbohydrates; Copyright Page; Contents; Contributors; Preface; Methods in Enzymology; Chapter 1: Introduction to Polyketide Biosynthesis; 1. Introduction; 1.1. Types of PKS; 1.2. Type II PKS; 1.3. Type III PKS; 1.4. Type I PKS; 1.5. Combinatorial biosynthesis: Prospects and progress; Acknowledgments; Chapter 2: Structural Enzymology of polyketide Synthases; Chapter 3: Chapter Three: Fungal Type I Polyketide Synthases; 1. Introduction

2. Partially Reducing PKSs: 6-Methylsalicylate Synthase3. Nonreducing PKSs; 3.1. Norsolorinic acid synthase; 3.2. Tetrahydroxynaphthalene synthase; 3.3. Bikaverin nonaketide synthase; 4. Highly Reducing PKSs; 4.1. Lovastatin (LNKS and LDKS); 4.2. HR PKS-NRPS: Fusarin and tenellin synthetases; 5. NR/HR PKS Hybrid Systems: Zearalenone (ZAE1 and ZAE2); 6. Conclusions; References; Chapter 4: Tandem Acyl Carrier Protein Domains in Polyunsaturated Fatty Acid Synthases; 1. Introduction; 2. Methods; 2.1. Production of PUFAs in E. coli by expressing the PUFAS genes

2.2. Mapping the active sites of PfaA-ACPs by site-directed mutagenesis2.3. Overproduction of each of the PfaA-ACPs; 2.4. Overproduction of PfaE and Svp PPTases; 2.5. In vivo and in vitro preparation of the holo-form of PfaA-ACPs; 2.6. Elucidation of the relationship between PUFA production and the number of active ACPs; 3. Conclusion; References; Chapter 5: Iterative Type I Polyketide Synthases for Enediyne Core Biosynthesis; 1. Introduction; 2. Methods; 2.1. PCR amplification of PKSE cassettes for predictive classification of new enediynes

2.2. Heterologous expression and overproduction of PKSE proteins2.3. Production and isolation of the polyene intermediate from 9-membered PKSEs; 2.4. Production of apo-ACPs from PKSE for in vitro functional analyses; 2.5. In vitro preparation of holo-ACPs; 3. Conclusion; Acknowledgments; References; Chapter 6: Chapter Six The DEBS Paradigm for Type I Modular Polyketide Synthases and Beyond; 1. Introduction; 2. DEBS and the Concept of a Module; 2.1. Generalizability of the DEBS paradigm; 3. Beyond the DEBS Paradigm; 3.1. Specificity of the AT domains; 3.2. Novel loading modules

3.3. Methylation domains3.4. Trans PKS activities; 3.5. Unusual modular organization; 3.6. Unusual module functions; 3.7. Intermodular interactions; 4. Conclusion; Chapter 7: Formation and Characterization of Acyl Carrier Protein-Linked Polyketide Synthase Extender Units; 1. Introduction; 2. Overproduction and Purification of Recombinant Proteins; 2.1. Principle; 2.2. Materials; 2.3. Heterologous overproduction of proteins; 2.4. Purification of enzymes using batch-binding method with Nickel-NTA resin; 3. Formation and Characterization of Hydroxymalonyl-ACP and Aminomalonyl-ACP

3.1. In vitro phosphopantetheinylation of the ACPs ZmaD and ZmaH

Notes:

Description based upon print version of record.

Includes bibliographical references and index.

ISBN:

9786612120947

9781282120945

1282120948

9780080923369

0080923364

OCLC:

435549930