The Library
Chapter 17 siderophore biosynthesis : a substrate specificity assay for nonribosomal peptide synthetase‐independent siderophore synthetases involving trapping of acyl‐adenylate intermediates with hydroxylamine
Tools
Tools
Tools
Kadi, Nadia and Challis, Gregory L. (2009) Chapter 17 siderophore biosynthesis : a substrate specificity assay for nonribosomal peptide synthetase‐independent siderophore synthetases involving trapping of acyl‐adenylate intermediates with hydroxylamine. Methods in Enzymology, Vol.458 . pp. 431-457. doi:10.1016/S0076-6879(09)04817-4 ISSN 0076-6879.
Research output not available from this repository.
Request-a-Copy directly from author or use local Library Get it For Me service.
Official URL: http://dx.doi.org/10.1016/S0076-6879(09)04817-4
Abstract
Siderophores are an important group of structurally diverse natural products that play key roles in ferric iron acquisition in most microorganisms. Two major pathways exist for siderophore biosynthesis. One is dependent on nonribosomal peptide synthetase (NRPS) multienzymes. The enzymology of several NRPS‐dependent pathways to structurally diverse siderophores has been intensively studied for more than 10 years and is generally well understood. The other major pathway is NRPS‐independent. It relies on a novel family of synthetase enzymes that until recently has received very little attention. Over the last 2 years, these enzymes have begun to be intensively investigated and several examples have now been characterized. In this article, we give an overview of the enzymology of NRPS‐dependent and NRPS‐independent pathways for siderophore biosynthesis, using selected examples to highlight key features.
An important facet of many studies of the enzymology of siderophore biosynthesis has been to investigate the substrate specificity of the synthetase enzymes involved. For NRPS‐dependent pathways, the ATP–pyrophophate exchange assay has been widely used to investigate the substrate specificity of adenylation domains within the synthetase multienzymes. This assay is ineffective for NRPS‐independent siderophore (NIS) synthetases, probably because pyrophosphate is not released from the enzyme after the carboxylic acid substrate and ATP react to form an acyl adenylate. An alternative assay for enzymes that form acyl adenylates involves trapping of the activated carboxyl group with hydroxylamine to form a hydroxamic acid that can be converted to its ferric complex and detected spectrophotometrically. This assay has not been widely used for NRPS adenylation domains. Here, we show that it is an effective assay for examining the carboxylic acid substrate specificity of NIS synthetases. Application of the assay to the type B NIS synthetase AcsA shows that it is selective for α‐ketoglutaric acid, confirming a bioinformatics‐based prediction of the substrate specificity of this enzyme.
| Item Type: | Journal Article | ||||
|---|---|---|---|---|---|
| Subjects: | Q Science > QD Chemistry | ||||
| Divisions: | Faculty of Science, Engineering and Medicine > Science > Chemistry | ||||
| Library of Congress Subject Headings (LCSH): | Siderophores -- Synthesis, Hydroxylamine, Ligases | ||||
| Journal or Publication Title: | Methods in Enzymology | ||||
| Publisher: | Academic Press | ||||
| ISSN: | 0076-6879 | ||||
| Book Title: | Complex Enzymes in Microbial Natural Product Biosynthesis, Part A: Overview Articles and Peptides | ||||
| Official Date: | 2009 | ||||
| Dates: |
|
||||
| Volume: | Vol.458 | ||||
| Page Range: | pp. 431-457 | ||||
| DOI: | 10.1016/S0076-6879(09)04817-4 | ||||
| Status: | Peer Reviewed | ||||
| Publication Status: | Published | ||||
| Access rights to Published version: | Restricted or Subscription Access | ||||
| Description: | Issue entitled: Complex Enzymes in Microbial Natural Product Biosynthesis, Part A: Overview Articles and Peptides |
Request changes or add full text files to a record
Repository staff actions (login required)
 |
View Item |
