Dataset Information

Structural characterization of new microcystins containing tryptophan and oxidized tryptophan residues.

ABSTRACT: Microcystins are cyclic peptides produced by cyanobacteria, which can be harmful to humans and animals when ingested. Eight of the (more than) 90 microcystin variants presently characterized, contain the amino acid tryptophan. The well-researched oxidation products of tryptophan; kynurenine, oxindolylalanine, and N-formylkynurenine, have been previously identified in intact polypeptides but microcystin congeners containing oxidized tryptophan moieties have not been reported. Liquid chromatography-tandem mass spectrometric analysis of an extract of Microcystis CAWBG11 led to the tentative identification of two new tryptophan-containing microcystins (MC?WAba and MC-WL), as well as eight other microcystin analogs containing kynurenine, oxindolylalanine and N?formylkynurenine (Nfk). Investigation of one of these congeners (MC?NfkA) by nuclear magnetic resonance spectroscopy was used to verify the presence of Nfk in the microcystin. Liquid chromatography-mass spectrometry analysis of a tryptophan oxidation experiment demonstrated that tryptophan-containing microcystins could be converted into oxidized tryptophan analogs and that low levels of oxidized tryptophan congeners were present intracellularly in CAWBG11. MC-NfkR and MC-LNfk were detected in standards of MC-WR and MC-LW, indicating that care during storage of tryptophan-containing microcystins is required.

SUBMITTER: Puddick J

PROVIDER: S-EPMC3766880 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Structural characterization of new microcystins containing tryptophan and oxidized tryptophan residues.

Puddick Jonathan J Prinsep Michèle R MR Wood Susanna A SA Miles Christopher O CO Rise Frode F Cary Stephen Craig SC Hamilton David P DP Wilkins Alistair L AL

Marine drugs 20130821 8

Microcystins are cyclic peptides produced by cyanobacteria, which can be harmful to humans and animals when ingested. Eight of the (more than) 90 microcystin variants presently characterized, contain the amino acid tryptophan. The well-researched oxidation products of tryptophan; kynurenine, oxindolylalanine, and N-formylkynurenine, have been previously identified in intact polypeptides but microcystin congeners containing oxidized tryptophan moieties have not been reported. Liquid chromatograph ...[more]

PMID: 23966035

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Project description:Hepatotoxic microcystins (MCs) are the most widespread class of cyanotoxins and the one that has most often been implicated in cyanobacterial toxicosis. One of the main challenges in studying and monitoring MCs is the great structural diversity within the class. The full chemical structure of the first MC was elucidated in the early 1980s and since then, the number of reported structural analogues has grown steadily and continues to do so, thanks largely to advances in analytical methodology. The structures of some of these analogues have been definitively elucidated after chemical isolation using a combination of techniques including nuclear magnetic resonance, amino acid analysis, and tandem mass spectrometry (MS/MS). Others have only been tentatively identified using liquid chromatography-MS/MS without chemical isolation. An understanding of the structural diversity of MCs, the genetic and environmental controls for this diversity and the impact of structure on toxicity are all essential to the ongoing study of MCs across several scientific disciplines. However, because of the diversity of MCs and the range of approaches that have been taken for characterizing them, comprehensive information on the state of knowledge in each of these areas can be challenging to gather. We have conducted an in-depth review of the literature surrounding the identification and toxicity of known MCs and present here a concise review of these topics. At present, at least 279 MCs have been reported and are tabulated here. Among these, about 20% (55 of 279) appear to be the result of chemical or biochemical transformations of MCs that can occur in the environment or during sample handling and extraction of cyanobacteria, including oxidation products, methyl esters, or post-biosynthetic metabolites. The toxicity of many MCs has also been studied using a range of different approaches and a great deal of variability can be observed between reported toxicities, even for the same congener. This review will help clarify the current state of knowledge on the structural diversity of MCs as a class and the impacts of structure on toxicity, as well as to identify gaps in knowledge that should be addressed in future research.

Project description:Tryptophan (Trp) is a naturally occurring amino acid, which exhibits fluorescence emission properties that are dependent on the polarity of the local environment around the Trp side chain. However, this sensitivity also complicates interpretation of fluorescence emission data. A non-natural analogue of tryptophan, β-(1-azulenyl)-L-alanine, exhibits fluorescence insensitive to local solvent polarity and does not impact the structure or characteristics of several peptides examined. In this study, we investigated the effect of replacing Trp with β-(1-azulenyl)-L-alanine in the well-known bee-venom peptide melittin. This peptide provides a model framework for investigating the impact of replacing Trp with β-(1-azulenyl)-L-alanine in a functional peptide system that undergoes significant shifts in Trp fluorescence emission upon binding to lipid bilayers. Microbiological methods including assessment of the antimicrobial activity by minimal inhibitory concentration (MIC) assays and bacterial membrane permeability assays indicated little difference between the Trp and the β-(1-azulenyl)-L-alanine-substituted versions of melittin. Circular dichroism spectroscopy showed both that peptides adopted the expected α-helical structures when bound to phospholipid bilayers and electrophysiological analysis indicated that both created membrane disruptions leading to significant conductance increases across model membranes. Both peptides exhibited a marked protection of the respective fluorophores when bound to bilayers indicating a similar membrane-bound topology. As expected, while fluorescence quenching and CD indicate the peptides are stably bound to lipid vesicles, the peptide containing β-(1-azulenyl)-L-alanine exhibited no fluorescence emission shift upon binding while the natural Trp exhibited >10 nm shift in emission spectrum barycenter. Taken together, the β-(1-azulenyl)-L-alanine can serve as a solvent insensitive alternative to Trp that does not have significant impacts on structure or function of membrane interacting peptides.

Dataset Information

Structural characterization of new microcystins containing tryptophan and oxidized tryptophan residues.

Publications

Structural characterization of new microcystins containing tryptophan and oxidized tryptophan residues.

Similar Datasets

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