An unusual cleavage reaction of a peptide observed during dithiotreitol and tris(2-carboxyethyl)phosphine reduction: application to sequencing of HpTx2 spider toxin using nanospray tandem mass spectrometry

A recombinant peptidic spider toxin, HpTx2, was investigated directly by nanoelectrospray tandem mass spectrometry (MS/MS). This 30‐residue toxin possesses a highly knotted structure with cystines arranged in close proximity. The low‐energy collision‐induced dissociation MS/MS spectrum of the [M+4H]...

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Bibliographic Details
Published in:Rapid communications in mass spectrometry 2004-01, Vol.18 (12), p.1317-1323
Main Authors: Legros, Christian, Célérier, Marie-Louise, Guette, Catherine
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Dithiothreitol - chemistry
Molecular Sequence Data
Nanotechnology
Oxidation-Reduction
Peptides - chemistry
Phosphines - chemistry
Sequence Analysis, Protein
Spectrometry, Mass, Electrospray Ionization - methods
Spider Venoms - chemistry
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Summary:A recombinant peptidic spider toxin, HpTx2, was investigated directly by nanoelectrospray tandem mass spectrometry (MS/MS). This 30‐residue toxin possesses a highly knotted structure with cystines arranged in close proximity. The low‐energy collision‐induced dissociation MS/MS spectrum of the [M+4H]4+ ion permitted characterization of the C‐terminal sequence of HpTx2 up to Cys26 that is involved in a disulfide bridge. Chemical pre‐treatment with DTT or TCEP was then investigated, and it was found that an unexpected cleavage reaction of HpTx2 gave two smaller peptides which were completely sequenced by MS/MS experiments using a Qq‐TOF mass spectrometer. This unusual hydrolysis reaction facilitated the determination of the complete sequence of the HpTx2 toxin. Copyright © 2004 John Wiley & Sons, Ltd.
ISSN:0951-4198
1097-0231
DOI:10.1002/rcm.1490