Detection and identification of antibacterial proteins in snake venoms using at-line nanofractionation coupled to LC-MS

Marija Mladic; Julien Slagboom; Jeroen Kool; Freek J. Vonk; Gilles P. van Wezel; Michael K. Richardson

doi:10.1016/j.toxicon.2018.08.009

Detection and identification of antibacterial proteins in snake venoms using at-line nanofractionation coupled to LC-MS

Marija Mladic, Julien Slagboom, Jeroen Kool, Freek J. Vonk, Gilles P. van Wezel, Michael K. Richardson^*

^*Corresponding author for this work

Research output: Contribution to Journal › Article › Academic › peer-review

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Abstract

This study describes the application of at-line nanofractionation to the screening of snake venoms for antibacterial activity against Gram-positive and Gram-negative bacteria, the detection of proteins of interest, and their partial or full identification. A method was developed to identify bioactive peptides in crude snake venoms based on reversed-phase liquid chromatography (LC), with parallel nanofractionation onto 384-well plates and mass spectrometry (MS). Bioactivity assays were based on a resazurin-reduction assay. Accurate masses of the bioactive peptides were determined, and peptides were then identified via nanoLC–MS/MS analysis of tryptic digests, allowing full or partial identification of the bioactive proteins. Crude venoms from 41 species were screened for their antibacterial bioactivity. Venoms showing the highest activity were further screened using at-line nanofractionation, which resulted in the elucidation of 28 bioactive proteins.

Original language	English
Pages (from-to)	66-74
Number of pages	9
Journal	Toxicon
Volume	155
Early online date	31 Aug 2018
DOIs	https://doi.org/10.1016/j.toxicon.2018.08.009
Publication status	Published - 1 Dec 2018

Funding

This work was supported by by grant nr 731.014.206 from the Netherlands Organization for Scientific Research (NWO) to M.K.R. and G.P.vW. Appendix A

Funders	Funder number
Netherlands Organization for Scientific Research
Nederlandse Organisatie voor Wetenschappelijk Onderzoek	731.014.206

Keywords

Antibacterial activity
Liquid chromatography-mass spectrometry
Resazurin-reduction assay
Snake venom

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Detection and identification of antibacterial proteins in snake venoms using at-line nanofractionation coupled to LC-MSFinal published version, 716 KBLicence: Article 25fa Dutch Copyright Act

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title = "Detection and identification of antibacterial proteins in snake venoms using at-line nanofractionation coupled to LC-MS",

abstract = "This study describes the application of at-line nanofractionation to the screening of snake venoms for antibacterial activity against Gram-positive and Gram-negative bacteria, the detection of proteins of interest, and their partial or full identification. A method was developed to identify bioactive peptides in crude snake venoms based on reversed-phase liquid chromatography (LC), with parallel nanofractionation onto 384-well plates and mass spectrometry (MS). Bioactivity assays were based on a resazurin-reduction assay. Accurate masses of the bioactive peptides were determined, and peptides were then identified via nanoLC–MS/MS analysis of tryptic digests, allowing full or partial identification of the bioactive proteins. Crude venoms from 41 species were screened for their antibacterial bioactivity. Venoms showing the highest activity were further screened using at-line nanofractionation, which resulted in the elucidation of 28 bioactive proteins.",

keywords = "Antibacterial activity, Liquid chromatography-mass spectrometry, Resazurin-reduction assay, Snake venom",

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AU - Mladic, Marija

AU - Slagboom, Julien

AU - Kool, Jeroen

AU - Vonk, Freek J.

AU - van Wezel, Gilles P.

AU - Richardson, Michael K.

PY - 2018/12/1

Y1 - 2018/12/1

N2 - This study describes the application of at-line nanofractionation to the screening of snake venoms for antibacterial activity against Gram-positive and Gram-negative bacteria, the detection of proteins of interest, and their partial or full identification. A method was developed to identify bioactive peptides in crude snake venoms based on reversed-phase liquid chromatography (LC), with parallel nanofractionation onto 384-well plates and mass spectrometry (MS). Bioactivity assays were based on a resazurin-reduction assay. Accurate masses of the bioactive peptides were determined, and peptides were then identified via nanoLC–MS/MS analysis of tryptic digests, allowing full or partial identification of the bioactive proteins. Crude venoms from 41 species were screened for their antibacterial bioactivity. Venoms showing the highest activity were further screened using at-line nanofractionation, which resulted in the elucidation of 28 bioactive proteins.

AB - This study describes the application of at-line nanofractionation to the screening of snake venoms for antibacterial activity against Gram-positive and Gram-negative bacteria, the detection of proteins of interest, and their partial or full identification. A method was developed to identify bioactive peptides in crude snake venoms based on reversed-phase liquid chromatography (LC), with parallel nanofractionation onto 384-well plates and mass spectrometry (MS). Bioactivity assays were based on a resazurin-reduction assay. Accurate masses of the bioactive peptides were determined, and peptides were then identified via nanoLC–MS/MS analysis of tryptic digests, allowing full or partial identification of the bioactive proteins. Crude venoms from 41 species were screened for their antibacterial bioactivity. Venoms showing the highest activity were further screened using at-line nanofractionation, which resulted in the elucidation of 28 bioactive proteins.

KW - Antibacterial activity

KW - Liquid chromatography-mass spectrometry

KW - Resazurin-reduction assay

KW - Snake venom

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Detection and identification of antibacterial proteins in snake venoms using at-line nanofractionation coupled to LC-MS

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Keywords

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