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Related Concept Videos

Peptide Identification Using Tandem Mass Spectrometry01:33

Peptide Identification Using Tandem Mass Spectrometry

Tandem mass spectrometry, also known as MS/MS or MS2, is an analytical technique that employs two mass analyzers. Essentially it is a series of mass spectrometers that helps isolate a particular biomolecule and then helps study its chemical properties.
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...

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Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
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Improved proteomic analysis pipeline for LC-ETD-MS/MS using charge enhancing methods.

Li-Qi Xie1, Cheng-Pin Shen, Min-Bo Liu

  • 1Shanghai Cancer Center and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, People's Republic of China. luhaojie@fudan.edu.cn

Molecular Biosystems
|July 21, 2012
PubMed
Summary

Electron transfer dissociation (ETD) in mass spectrometry struggles with low scores for 2+ ions. Combining ion charge enhancement (like m-NBA) with search algorithms (X!Tandem, pFind) significantly boosts peptide identification rates.

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Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
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Published on: February 27, 2020

Area of Science:

  • Mass Spectrometry
  • Proteomics
  • Analytical Chemistry

Background:

  • Electron transfer dissociation (ETD) is a valuable peptide fragmentation technique.
  • ETD spectra often yield lower scores for 2+ ions, hindering identification.
  • Optimizing ETD for complex samples requires enhanced strategies.

Purpose of the Study:

  • To systematically evaluate methods for improving ETD peptide identification.
  • To investigate the combined effects of ion charge enhancement and search algorithms.
  • To establish optimal protocols for high-throughput proteomics.

Main Methods:

  • Applied ion charge enhancing methods: dimethylation, guanidination, m-nitrobenzyl alcohol (m-NBA) adduction, and Lys-C digestion.
  • Utilized differential search algorithms: Mascot, Sequest, OMSSA, pFind, and X!Tandem.
  • Benchmarked performance using a simple protein (BSA) and a complex cell lysate (AMJ2).

Main Results:

  • X!Tandem and pFind identified 92.65% of spectra in AMJ2 cell lines.
  • m-NBA adduction increased average charge state by 5-10% and identification numbers.
  • Lys-C digestion predominantly produced triple-charged peptides.

Conclusions:

  • Combining m-NBA and Lys-C strategies with X!Tandem and pFind significantly enhances ETD identification.
  • Differential search algorithms and ion charge enhancement offer complementary benefits.
  • Optimized ETD protocols are crucial for deep proteome coverage.