Fourier-transform mass spectrometry for automated fragmentation and identification of 5-20 kDa proteins in mixtures

Jeffrey R. Johnson, Fanyu Meng, Andrew J. Forbes, Benjamin J. Cargile, Neil L. Kelleher

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

When presented with a mixture of intact proteins, electrospray ionization with Fourier-transform mass spectrometry (ESI-FTMS) has the capability to obtain direct fragmentation information from isolated ions. However, the automation of this capability has not been achieved to date. We have developed software for unattended acquisition of protein tandem mass spectrometry (MS/MS) data and batch processing of the resulting files for identification of whole proteins. Mixtures of both protein standards (8-29 kDa) and Methanococcus jannaschii cytosolic proteins (up to six components < 20 kDa) were infused via an autosampler, and MS/MS data were acquired without human intervention. The acquisition software recognizes ESI charge state patterns, generates protein-specific isolation waveforms on-the-fly, and fragments ions using two different infrared laser times. In addition to protein standards, five wild-type proteins (7-14 kDa) were identified automatically with 100% sequence coverage from the M. jannaschii database. The software underpins a measurement platform for sample-dependent acquisition of MS/MS data for whole proteins, a critical step to realize proteomics with 100% sequence coverage in a higher throughput setting.

Original languageEnglish (US)
Pages (from-to)3217-3223
Number of pages7
JournalELECTROPHORESIS
Volume23
Issue number18
DOIs
StatePublished - Sep 2002

Keywords

  • Automation
  • Electrospray
  • Fourier-transform mass spectrometry
  • Intact proteins
  • Tandem mass spectrometry

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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