Combining [13C6]-phenylisothiocyanate and the edman degradation reaction: A possible breakthrough for absolute quantitative proteomics together with protein identification

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Abstract

This manuscript describes the results of a preliminary experiment performed as 'proof of concept' of a novel approach to absolute quantitation of proteins without the use of standard proteins. Absolute quantitation remains a challenging issue in the proteomics field. Therefore, we propose a combination of [13C6]-phenylisothiocyanate (PITC) and the Edman degradation reaction as a possible breakthrough. [13C6]-PITC was synthesized from [13C6]-aniline with O,O(-di-2-pyridyl thiocarbonate to prepare [13C6]-phenylthiohydantoin (PTH)-amino acids as internal standards. Upon the Edman degradation reaction, it has been confirmed that a model protein, bovine serum albumin (BSA), releases the N-terminal amino acid quantitatively as PTH-Asp. The standard curve of PTH-Asp against [13C6]-PTH-Asp showed good linearity (r2 1/40. 9977). BSA could be quantified as PTH-Asp using the standard curve. In addition, the residual des-Asp1-BSA provided sufficient information for further protein identification.

Original languageEnglish
Pages (from-to)173-179
Number of pages7
JournalRapid Communications in Mass Spectrometry
Volume24
Issue number2
DOIs
Publication statusPublished - 2010 Jan 30

ASJC Scopus subject areas

  • Analytical Chemistry
  • Spectroscopy
  • Organic Chemistry

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