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Article (Scientific journals)
New method for characterizing highly disulfide-bridged peptides in complex mixtures: Application to toxin identification from crude venoms
Quinton, Loïc; Demeure, Kevin; Dobson, Rowan et al.
2007In Journal of Proteome Research, 6 (8), p. 3216-3223
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Keywords :
in-source decay; post-source decay; conotoxin; snake toxins; disulfide-bridged peptides; top-down sequencing; mass spectrometry; venom
Abstract :
[en] Animal venoms are highly complex mixtures that can contain many disulfide-bridged toxins. This work presents an LC-MALDI approach allowing (1) a rapid classification of toxins according to their number of disulfide bonds and (2) a rapid top-down sequencing of the toxins using a new MALDI matrix enhancing in-source decay (ISD). The crude venom is separated twice by LC: the fractions of the first separation are spotted on the MALDI matrix alpha-cyano-4-hydroxycinnamic acid (CHCA) and the others using 1,5-diaminonaphthalene (1,5-DAN). CHCA spots are more convenient for obtaining a precise mass fingerprint of a large number of peptides; however, the analysis of 1,5-DAN spots allows the number of disulfide bridges to be counted owing to their partial in-plume reduction by this particular matrix. Subsequently, the disulfide bonds of all peptides present in the crude venom were reduced by an excess of tris(carboxyethyl)phosphine before the LC separation and were subjected to the same analysis in CHCA and 1,5-DAN. Toxins were sequenced using a TOF/TOF analysis of metastable fragments from CHCA spots and ISD fragmentation from 1,5-DAN spots. Novel conotoxin sequences were found using this approach. The use of 1,5-DAN for ISID top-down sequencing is also illustrated for higher molecular weight toxins such as snake cardiotoxins and neurotoxins (>6500 Da), where sequence coverage >70% is obtained from the c-ion series.
Research center :
Giga-Systems Biology and Chemical Biology - ULiège
Disciplines :
Chemistry
Author, co-author :
Quinton, Loïc  ;  Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Demeure, Kevin ;  Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Dobson, Rowan ;  Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Gilles, Nicolas;  CEA/DSV/iBiTec-S/SIMOPRO, Gif-sur-Yvette Cedex 91191, France
Gabelica, Valérie ;  Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
De Pauw, Edwin  ;  Université de Liège - ULiège > Département de chimie (sciences) > Chimie physique, spectrométrie de masse
Language :
English
Title :
New method for characterizing highly disulfide-bridged peptides in complex mixtures: Application to toxin identification from crude venoms
Publication date :
2007
Journal title :
Journal of Proteome Research
ISSN :
1535-3893
eISSN :
1535-3907
Publisher :
Amer Chemical Soc, Washington, United States - Washington
Volume :
6
Issue :
8
Pages :
3216-3223
Peer reviewed :
Peer Reviewed verified by ORBi
Funders :
F.R.S.-FNRS - Fonds de la Recherche Scientifique [BE]
Available on ORBi :
since 26 November 2008

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