Rapid screening and identification of ACE inhibitors in snake venoms using at-line nanofractionation LC-MS View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2017-10

AUTHORS

Marija Mladic, Tessa de Waal, Lindsey Burggraaff, Julien Slagboom, Govert W. Somsen, Wilfried M. A. Niessen, R. Manjunatha Kini, Jeroen Kool

ABSTRACT

This study presents an analytical method for the screening of snake venoms for inhibitors of the angiotensin-converting enzyme (ACE) and a strategy for their rapid identification. The method is based on an at-line nanofractionation approach, which combines liquid chromatography (LC), mass spectrometry (MS), and pharmacology in one platform. After initial LC separation of a crude venom, a post-column flow split is introduced enabling parallel MS identification and high-resolution fractionation onto 384-well plates. The plates are subsequently freeze-dried and used in a fluorescence-based ACE activity assay to determine the ability of the nanofractions to inhibit ACE activity. Once the bioactive wells are identified, the parallel MS data reveals the masses corresponding to the activities found. Narrowing down of possible bioactive candidates is provided by comparison of bioactivity profiles after reversed-phase liquid chromatography (RPLC) and after hydrophilic interaction chromatography (HILIC) of a crude venom. Additional nanoLC-MS/MS analysis is performed on the content of the bioactive nanofractions to determine peptide sequences. The method described was optimized, evaluated, and successfully applied for screening of 30 snake venoms for the presence of ACE inhibitors. As a result, two new bioactive peptides were identified: pELWPRPHVPP in Crotalus viridis viridis venom with IC50 = 1.1 μM and pEWPPWPPRPPIPP in Cerastes cerastes cerastes venom with IC50 = 3.5 μM. The identified peptides possess a high sequence similarity to other bradykinin-potentiating peptides (BPPs), which are known ACE inhibitors found in snake venoms. More... »

PAGES

5987-5997

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/s00216-017-0531-3

DOI

http://dx.doi.org/10.1007/s00216-017-0531-3

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1091146557

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/28801827


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