Conformational properties of

a-dendrotoxin using electrospray mass spectrometry

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Eur. J. Mass Spectrom. 7, 373 - 383 (2001)

Conformational properties of a-dendrotoxin using electrospray mass spectrometry

Hélène Belva and Catherine Lange^*
Spectrométrie de Masse Bio-Organique, CNRS-UMR 6014, INSERM-IFR23, UFR des Sciences, Université de Rouen, 76821 Mont Saint Aignan-Cedex, France. E-mail:
Isabelle Segalas
RMN Modélisation Moléculaire, CNRS-UMR 6014, INSERM-IFR23, UFR des Sciences, Université de Rouen, 76821 Mont Saint Aignan-Cedex, France

ABSTRACT:

To examine the conformation of a-dendrotoxin, charge state distributions (CSD) and hydrogen/deuterium (H/D) isotopic exchange using electrospray-ionization mass spectrometry [ES-MS] have been used. Here, it cannot be deduced from the CSDs that some basic residues are hidden in the core of the structure. At neutral pH, the CSDs reflect the equilibrium bulk solution exactly. Seven basic residues seem to be shielded from the solvent, according to calculations from the X-ray structure. But, under acidic conditions, the CSDs observed in mass spectra are not a true reflection of the equilibrium bulk solution, partly because of an increased intramolecular electrostatic effect with the gaseous ions. H/D exchange experiments show that three to seven hydrogens are involved in hydrogen bonds. The resistance to urea denaturation has also been investigated. Urea and 1,4-dithiothreitol (DTT) denaturation together lead to unfolded conformers. During this reduction of the bridges, it is observed that the Cys16Cys40 and Cys32Cys53 linkages are more accessible to the reducing agent than the Cys7Cys57 linkage, which is very important by maintaining a compact fold.

Keywords: a-dendrotoxin, electrospray mass spectrometry, multiple disulfide bridges, basic peptide, charge state distributions, hydrogen/deuterium exchange

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