Reference : Disulfide bond assignment and folding characterization of peptide toxins by Ion Mobil...
Scientific congresses and symposiums : Poster
Physical, chemical, mathematical & earth Sciences : Chemistry
http://hdl.handle.net/2268/93335
Disulfide bond assignment and folding characterization of peptide toxins by Ion Mobility Mass Spectrometry
English
Echterbille, Julien mailto [Université de Liège - ULg > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.) >]
Quinton, Loïc mailto [Université de Liège - ULg > Département de chimie (sciences) > Chimie biologique >]
Rosu, Frédéric [Université de Liège - ULg > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.) >]
De Pauw, Edwin mailto [Université de Liège - ULg > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.) >]
2011
Yes
No
International
59th Conference on Mass Spectrometry and Allied Topics
du 5 juin 2011 au 9 juin 2011
American Society for Mass Spectrometry
Denver
CO, USA
[en] Ion Mobility ; Disulfide bond ; Peptide toxins
[en] Introduction
Animal venoms are mainly composed of peptide toxins, which are highly structured by several disulfide bridges. Disulfide bridges are a key feature as (i) they increase the toxins efficiency by lowering their immunogenicity; (ii) they provide the adequate conformation for high affinity binding to the biological receptor. The sequencing and the determination of the cysteine pairing is still challenging and therefore an important step in their structure analysis and the understanding of their interactions with receptors. In this work, we present a new strategy to sequence structured toxins and assign S-S bridges using ion mobility resolved MS/MS.
Methods
The method relies on the analysis of partially reduced multiple-disulfide peptide. The mixture of the different forms is resolved by ion mobility, followed by MS/MS acquisition on each mobility separated species. The proof of concept has been successfully conducted on α-CnI, a toxin purified from the venom of Conus consors marine snail. The toxin sequence is GRCCHPACGKYYSC-NH2. It contains four cysteines linked together with two disulfide bridges. α-CnI was partially reduced by a small excess of tris(carboxyethyl)phosphine (10:1) at 56°C during 30min. The resulting mixture was purified by ZipTip C18 micro columns before analysis by infusion nanoESI-Synapt-G2. Fragmentation was performed after the mobility cell, to obtain specific fragments of each species. Mobilograms and mass spectra were analyzed using MassLynx (v4.1) and Driftscope (v2.1) from Waters.
Preliminary data
Partial reduction of a-CnI was performed in order to obtain a mixture of oxidized (the two disulfides are formed), reduced (the two disulfides have been reduced) and partially reduced forms (only one of the two disulfides has been reduced). The arrival time distribution of triply charged ions reveals the presence of 4 different species, characterized by a different relative cross sections in the gas-phase. The charge state of the ions influences the ion mobility separation. Mass matching allows identifying the species: the first mobility (the most compact structure) was identified to be the oxidized folded toxin (M=1541.58 Da). The latest peak, corresponding to the larger cross-section, was identified as the fully reduced toxin (M=1545.6 Da). The second and the third mobility peaks were attributed to the two partially reduced forms in which only one disulfide bridge was reduced (M=1543.59 Da). The change in ion mobility depends on which S-S bridge is reduced. Ion mobility separated species give characteristic fragment ions upon fragmentation in the transfer cell (i.e. after ion mobility separator). Interestingly, fragment ions coming from partially reduced species, especially the C-S or S-S bond cleavages, clearly indicates that the disulfide linkage of α-CnI is (Cys1-Cys3) and (Cys2-Cys4) as expected from literature. The method is now being applied with success to more complex systems containing 3 or 4 disulfide bridges. The influence of the charge state on the mobility separation is systematically analyzed in terms of structural implications.
Novel aspect
Sequencing and disulfide bridges assignment of peptide toxins using ion mobility resolved MS/MS
Laboratoire de Spectrométrie de Masse
Patrimoine ULg
http://hdl.handle.net/2268/93335

File(s) associated to this reference

Fulltext file(s):

FileCommentaryVersionSizeAccess
Restricted access
Poster_ASMS_IMMS.pdfAuthor postprint230.78 kBRequest copy

Bookmark and Share SFX Query

All documents in ORBi are protected by a user license.