Algorithm of Protein Sequence Determination by Combination of the Edman Degradation Method and Shotgun Mass Spectrometry

  • V.S. Skvortsov Institute of Biomedical Chemistry, 10 Pogodinskaya str., Moscow, 119121 Russia
  • A.V. Mikurova Institute of Biomedical Chemistry, 10 Pogodinskaya str., Moscow, 119121 Russia
  • N.E. Vavilov Institute of Biomedical Chemistry, 10 Pogodinskaya str., Moscow, 119121 Russia
  • V.G. Zgoda Institute of Biomedical Chemistry, 10 Pogodinskaya str., Moscow, 119121 Russia
Keywords: Edman degradation; shotgun mass spectrometry; de novo sequencing

Abstract

An algorithm combining advantages of the Edman degradation method and de novo mass-spectrometric sequencing was developed. The protein from the “Diaskintest” diagnostic test was used for analysis. The protein was digested with trypsin and 5 steps of Edman degradation were carried out sequentially for the mixture of peptides. At each stage, the resulting mixture was analyzed by shotgun mass spectrometry analysis. The results of mass-spectrometry were analyzed both by the well-known de novo sequencing programs Novor and PepNovo+, and by own program that clustered individual spectra with a C-terminal signature formed by Y-ions. This approach allows us to determine confidently the amino acid sequence of the N-terminal part of the peptides obtained after the protein hydrolysis by trypsin.

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Published
2018-12-25
Section
PROTOCOLS OF EXPERIMENTS, USEFUL MODELS, PROGRAMS AND SERVICES