Mass Spectrometry Characterization of the Thermal Decomposition/Digestion (TDD) at Cysteine in Peptides and Proteins in the Condensed Phase |
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Authors: | Franco Basile Shaofeng Zhang Sujit Kumar Kandar Liang Lu |
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Institution: | (1) Department of Chemistry, University of Wyoming, 1000 University Ave., Laramie, WY 82071, USA |
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Abstract: | We report on the characterization by mass spectrometry (MS) of a rapid, reagentless and site-specific cleavage at the N-terminus
of the amino acid cysteine (C) in peptides and proteins induced by the thermal decomposition at 220–250 °C for 10 s in solid
samples. This thermally induced cleavage at C occurs under the same conditions and simultaneously to our previously reported
thermally induced site-specific cleavage at the C-terminus of aspartic acid (D) (Zhang, S.; Basile, F. J. Proteome Res.
2007, 6, (5), 1700–1704). The C cleavage proceeds through cleavage of the nitrogen and α–carbon bond (N-terminus) of cysteine and
produces modifications at the cleavage site with an amidation (−1 Da) of the N-terminal thermal decomposition product and
a −32 Da mass change of the C-terminal thermal decomposition product, the latter yielding either an alanine or β-alanine residue
at the N-terminus site. These modifications were confirmed by off-line thermal decomposition electrospray ionization (ESI)-MS,
tandem MS (MS/MS) analyses and accurate mass measurements of standard peptides. Molecular oxygen was found to be required
for the thermal decomposition and cleavage at C as it induced an initial cysteine thiol side chain oxidation to sulfinic acid.
Similar to the thermally induced D cleavage, missed cleavages at C were also observed. The combined thermally induced digestion
process at D and C, termed thermal decomposition/digestion (TDD), was observed on several model proteins tested under ambient
conditions and the site-specificity of the method confirmed by MS/MS. |
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