Dataset Information

Constructing a Tandem Mass Spectral Library for Forensic Ricin Identification

ABSTRACT: Ricin, a protein found in castor seeds, is a lethal toxin that is designated as a category 2 select agent. Because castor seeds are easy to obtain and the toxin can be easily extracted, cases of attempted ricin poisoning are relatively common. A shotgun proteomics method for ricin identification has recently been developed (manuscript in preparation), in which ricin peptides are identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) followed by proteomics database search, and peptide-spectrum matches are verified and compared to standard spectra by a human expert. To make this process more reproducible, objective, and high-throughput, we have created a ricin spectral library for peptide identification to supplement the human review step. To construct these spectral libraries, two pure ricin samples (from a proposed standard reference material) and crude castor seed extracts were digested with trypsin and analyzed using a standard shotgun LC-MS/MS protocol. Spectral libraries were created from the filtered search results from four database search tools. The library was then used in a search using SpectraST on samples from castor seeds. Analysis showed that the spectral library search resulted in more peptides identified from crude castor seed samples compared to MS-GF+ and Sequest plus Percolator. These results suggest that computational comparison of putative ricin peptide spectra to library spectra can be an effective method of confirming the presence of ricin, and that spectral library search may be suitable to augment the more manual and subjective aspects of the currently recommended human expert review.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Ricinus Communis

SUBMITTER: Isabelle O'Bryon

LAB HEAD: Karen Wahl

PROVIDER: PXD013711 | Pride | 2019-10-02

REPOSITORIES: pride

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Dataset's files

Source:

			Action	DRS
	Aggregate_functions_script.r	Other
	AggregateandcreateCastorlibrary.Rmd	Other
	Aggregateandcreatericinlibrary.Rmd	Other
	Castorseedanalysis.Rmd	Other
	ComparericinPSMstoSSMs.Rmd	Other

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Publications

Constructing a Tandem Mass Spectral Library for Forensic Ricin Identification.

O'Bryon Isabelle I Tucker Abigail E AE Kaiser Brooke L D BLD Wahl Karen L KL Merkley Eric D ED

Journal of proteome research 20191014 11

Ricin, a protein found in castor seeds, is a lethal toxin that is designated as a category 2 select agent, and cases of attempted ricin poisoning are relatively common. Many methods to detect protein toxins such as ricin use targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify toxin peptides, usually tryptic peptides. The successful use of untargeted methods has also been reported. However, the use of untargeted proteomics methods, including database search, for peptide ...[more]

PMID: 31566388

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Project description:The toxin ricin has been shown to cause inflammatory lung damage, leading to pulmonary oedema and, at higher doses, mortality. In order to understand the genetic basis of this inflammatory cascade a custom microarray platform (1509 genes) directed towards immune and inflammatory markers was used to investigate the temporal expression profiles of genes in a Balb/c mouse model of inhalational ricin exposure. To facilitate examination of those genes involved in both inflammatory cascades and wound repair the dose which was investigated was sub-lethal across a 96 hour time course. Histopathology of the lung was mapped across the time course and genetic responses considered in the context of overall lung pathology. 685 genes were found to be statistically significantly different compared to controls, across the time course and these genes have been investigated in the context of their biological function in ricin poisoning. As well as confirming key inflammatory markers associated with ricin intoxication (TNFa and Il1b) several pathways that are altered in expression were identified following pulmonary exposure to ricin. These genes included those involved in cytokine-cytokine signalling cascades (IL1, IL1r, IL1r2, Ccl 4, 6, 10), focal adhesion (Fn1, ICAM1) and tissue remodelling (VEGF, Pim1). Furthermore, the observed alteration in expression of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) indicates a key role in membrane integrity and cellular adhesion in ricin poisoning. Data captured using this transcriptomic approach could be used to develop a specific approach to the treatment of inhalational ricin exposure. This work was conducted as part of a wider programme of work to compare a number of militarily relevant lung damaging agents, with a view to establishing a rational basis for the identification of more generic medical countermeasures. Four groups of six mice were exposed to ricin and a further groups exposed to vehicle only control (PBS). Following exposure mice were culled at 6 timepoints (1, 4, 7, 24, 48 and 96h). RNA was extracted and run on the custom array.