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Liquid chromatography electron capture dissociation tandem mass spectrometry (LC-ECD-MS/MS) versus liquid chromatography collision-induced dissociation tandem mass spectrometry (LC-CID-MS/MS) for the identification of proteins.

Creese AJ, Cooper HJ - J. Am. Soc. Mass Spectrom. (2007)

Bottom Line: Experiments were performed on a hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer.Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin, lysozyme, cytochrome c, alcohol dehydrogenase, and beta-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides.The results show that although protein coverage was lower for LC-ECD-MS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags, providing greater confidence in protein assignment.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham, United Kingdom.

ABSTRACT
Electron capture dissociation (ECD) offers many advantages over the more traditional fragmentation techniques for the analysis of peptides and proteins, although the question remains: How suitable is ECD for incorporation within proteomic strategies for the identification of proteins? Here, we compare LC-ECD-MS/MS and LC-CID-MS/MS as techniques for the identification of proteins. Experiments were performed on a hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer. Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin, lysozyme, cytochrome c, alcohol dehydrogenase, and beta-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides. The results show that although protein coverage was lower for LC-ECD-MS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags, providing greater confidence in protein assignment.

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Protein sequence coverage obtained for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS analyses. Mean values obtained over the six proteins are also shown.
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fig2: Protein sequence coverage obtained for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS analyses. Mean values obtained over the six proteins are also shown.

Mentions: Figure 2 shows the average overall protein sequence coverage for the six proteins. The values are calculated from the Bioworks searches. It clearly shows that generally LC-CID-MS/MS results in greater sequence coverage than LC-ECD-MS/MS: 24% versus 20%, respectively. There are variations for each protein: LC-CID-MS/MS results in greater sequence coverage for lysozyme, cytochrome c, alcohol dehydrogenase, and β-galactosidase, whereas the reverse was true for BSA and transferrin. For lysozyme, the average of three ECD analyses gave 24%, with the CID analyses resulting in 27% coverage. For cytochrome c, sequence coverages of 34% (ECD) and 39% (CID) were obtained. For alcohol dehydrogenase, ECD analyses gave an average of 5% sequence coverage, with the CID analyses producing 22% coverage. The average sequence coverage obtained for BSA was 24% (ECD) and 20% (CID), for apo-transferrin was 31% (ECD) and 21% (CID), and for β-galactosidase was 4% (ECD) and 14% (CID).


Liquid chromatography electron capture dissociation tandem mass spectrometry (LC-ECD-MS/MS) versus liquid chromatography collision-induced dissociation tandem mass spectrometry (LC-CID-MS/MS) for the identification of proteins.

Creese AJ, Cooper HJ - J. Am. Soc. Mass Spectrom. (2007)

Protein sequence coverage obtained for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS analyses. Mean values obtained over the six proteins are also shown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2572008&req=5

fig2: Protein sequence coverage obtained for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS analyses. Mean values obtained over the six proteins are also shown.
Mentions: Figure 2 shows the average overall protein sequence coverage for the six proteins. The values are calculated from the Bioworks searches. It clearly shows that generally LC-CID-MS/MS results in greater sequence coverage than LC-ECD-MS/MS: 24% versus 20%, respectively. There are variations for each protein: LC-CID-MS/MS results in greater sequence coverage for lysozyme, cytochrome c, alcohol dehydrogenase, and β-galactosidase, whereas the reverse was true for BSA and transferrin. For lysozyme, the average of three ECD analyses gave 24%, with the CID analyses resulting in 27% coverage. For cytochrome c, sequence coverages of 34% (ECD) and 39% (CID) were obtained. For alcohol dehydrogenase, ECD analyses gave an average of 5% sequence coverage, with the CID analyses producing 22% coverage. The average sequence coverage obtained for BSA was 24% (ECD) and 20% (CID), for apo-transferrin was 31% (ECD) and 21% (CID), and for β-galactosidase was 4% (ECD) and 14% (CID).

Bottom Line: Experiments were performed on a hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer.Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin, lysozyme, cytochrome c, alcohol dehydrogenase, and beta-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides.The results show that although protein coverage was lower for LC-ECD-MS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags, providing greater confidence in protein assignment.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham, United Kingdom.

ABSTRACT
Electron capture dissociation (ECD) offers many advantages over the more traditional fragmentation techniques for the analysis of peptides and proteins, although the question remains: How suitable is ECD for incorporation within proteomic strategies for the identification of proteins? Here, we compare LC-ECD-MS/MS and LC-CID-MS/MS as techniques for the identification of proteins. Experiments were performed on a hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer. Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin, lysozyme, cytochrome c, alcohol dehydrogenase, and beta-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides. The results show that although protein coverage was lower for LC-ECD-MS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags, providing greater confidence in protein assignment.

Show MeSH
Related in: MedlinePlus