Algorithm-assisted elucidation of disulfide structure: Application of the negative signature mass algorithm to mass-mapping the disulfide structure of the 12-cysteine transforming growth factor β type II receptor extracellular domain

Chad R. Borges, Jianfeng Qi, Wei Wu, Eric Torng, Andrew P. Hinck, Jack Throck Watson

Research output: Contribution to journalArticlepeer-review

10 Scopus citations

Abstract

The power of an algorithm-driven method for interpreting disulfide mass-mapping data is demonstrated in the context of determining the disulfide structure of the extracellular domain of the transforming growth factor β type II receptor, a 14-kDa cystinyl protein containing 12 cysteines in the form of six disulfide bonds. The disulfide mass-mapping methodology is based on partial reduction and cyanylation-induced cleavage of the cystinyl protein. Because the multiplicity of possible disulfide structures that must be considered grows rapidly with the number of cysteines, as does the difficulty in physically isolating each of the partially reduced and cyanylated isoforms of the analyte, manual data interpretation for disulfide mapping a cystinyl protein containing more than eight cysteines becomes unmanageable. Recently, we introduced the concept of a "negative signature mass algorithm" (NSMA) to determine the disulfide structure of a cystinyl protein by processing an input of its amino acid sequence and mass spectral data from analysis of its associated cyanylation-induced cleavage products. Here, we present experimental results to validate the NSMA concept. A key advantage of the NSMA, in addition to convenience and automation, is its capacity to interpret mass spectra from mixtures of cyanylation-induced cleavage fragments without separating the partially reduced isoforms of the cystinyl protein and without knowledge of the extent of partial reduction.

Original languageEnglish (US)
Pages (from-to)91-103
Number of pages13
JournalAnalytical Biochemistry
Volume329
Issue number1
DOIs
StatePublished - Jun 1 2004

Keywords

  • Cyanylation
  • Data processing
  • Disulfide bonds
  • Mass spectra
  • Negative signature mass
  • Partial reduction
  • Polypeptide cleavage
  • Transforming growth factor β

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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