High-throughput peptide identification from protein digests using data-dependent multiplexed tandem FTICR mass spectrometry coupled with capillary liquid chromatography

L. Li, C. D. Masselon, G. A. Anderson, L. Pasǎa-Tolić, Sang-Won Lee, Y. Shen, R. Zhao, M. S. Lipton, T. P. Conrads, N. Tolić, R. D. Smith

Research output: Contribution to journalArticle

84 Citations (Scopus)

Abstract

Tandem mass spectrometry (MS/MS) plays an important role in the unambiguous identification and structural elucidation of biomolecules. In contrast to conventional MS/MS approaches for protein identification where an individual polypeptide is sequentially selected and dissociated, a multiplexed-MS/MS approach increases throughput by selecting several peptides for simultaneous dissociation using either infrared multiphoton dissociation (IRMPD) or multiple frequency sustained off-resonance irradiation (SORI) collisionally induced dissociation (CID). The high mass measurement accuracy and resolution of FTICR combined with knowledge of peptide dissociation pathways allows the fragments arising from several different parent ions to be assigned. Herein we report the application of multiplexed-MS/MS coupled with on-line separations for the identification of peptides present in complex mixtures (i.e., whole cell lysate digests). Software was developed to enable "on-the-fly" data-dependent peak selection of a subset of polypeptides from each FTICR MS acquisition. In the subsequent MS/MS acquisitions, several coeluting peptides were fragmented simultaneously using either IRMPD or SORI-CID techniques. The utility of this approach has been demonstrated using a bovine serum albumin tryptic digest separated by capillary LC where multiple peptides were readily identified in single MS/MS acquisitions. We also present initial results from multiplexed-MS/MS analysis of a D. radiodurans whole cell digest to illustrate the utility of this approach for high-throughput analysis of a bacterial proteome.

Original languageEnglish
Pages (from-to)3312-3322
Number of pages11
JournalAnalytical Chemistry
Volume73
Issue number14
DOIs
Publication statusPublished - 2001 Jul 15
Externally publishedYes

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Liquid chromatography
Mass spectrometry
Throughput
Peptides
Proteins
Irradiation
Infrared radiation
Biomolecules
Proteome
Bovine Serum Albumin
Complex Mixtures
Ions

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

High-throughput peptide identification from protein digests using data-dependent multiplexed tandem FTICR mass spectrometry coupled with capillary liquid chromatography. / Li, L.; Masselon, C. D.; Anderson, G. A.; Pasǎa-Tolić, L.; Lee, Sang-Won; Shen, Y.; Zhao, R.; Lipton, M. S.; Conrads, T. P.; Tolić, N.; Smith, R. D.

In: Analytical Chemistry, Vol. 73, No. 14, 15.07.2001, p. 3312-3322.

Research output: Contribution to journalArticle

Li, L, Masselon, CD, Anderson, GA, Pasǎa-Tolić, L, Lee, S-W, Shen, Y, Zhao, R, Lipton, MS, Conrads, TP, Tolić, N & Smith, RD 2001, 'High-throughput peptide identification from protein digests using data-dependent multiplexed tandem FTICR mass spectrometry coupled with capillary liquid chromatography', Analytical Chemistry, vol. 73, no. 14, pp. 3312-3322. https://doi.org/10.1021/ac010192w
Li, L. ; Masselon, C. D. ; Anderson, G. A. ; Pasǎa-Tolić, L. ; Lee, Sang-Won ; Shen, Y. ; Zhao, R. ; Lipton, M. S. ; Conrads, T. P. ; Tolić, N. ; Smith, R. D. / High-throughput peptide identification from protein digests using data-dependent multiplexed tandem FTICR mass spectrometry coupled with capillary liquid chromatography. In: Analytical Chemistry. 2001 ; Vol. 73, No. 14. pp. 3312-3322.
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