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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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Proportion of peptides identified containing sequence tags of ≥6 consecutive amino acids for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS. Mean values obtained over the six proteins are also shown.
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fig4: Proportion of peptides identified containing sequence tags of ≥6 consecutive amino acids for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS. Mean values obtained over the six proteins are also shown.

Mentions: Figure 4 shows the percentage of peptides detected for each analysis type that had ≥6 amino acid sequence tags. An average of 62.0% of the lysozyme peptides detected in the LC-ECD-MS/MS analyses had sequence tags of ≥6 amino acids compared with 40.8% of those detected in the LC-CID-MS/MS analyses. The results for the remaining proteins were: cytochrome c 62.2% (ECD) and 18.9% (CID); alcohol dehydrogenase 18% (ECD) and 27.2% (CID); BSA 35.6% (ECD) and 22.2% (CID); apo-transferrin 53.3% (ECD) and 22.9% (CID); and β-galactosidase 0% (ECD) and 11.0% (CID). These results suggest that LC-ECD-MS/MS generally produces longer and therefore more reliable sequence tags than LC-CID-MS/MS. When all six proteins are considered, the ECD analyses resulted in 38.5% coverage, whereas the CID analyses gave 23.8% coverage.


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)

Proportion of peptides identified containing sequence tags of ≥6 consecutive amino acids for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS. 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

fig4: Proportion of peptides identified containing sequence tags of ≥6 consecutive amino acids for the six proteins (averaged over three repeats) for LC-ECD-MS/MS and LC-CID-MS/MS. Mean values obtained over the six proteins are also shown.
Mentions: Figure 4 shows the percentage of peptides detected for each analysis type that had ≥6 amino acid sequence tags. An average of 62.0% of the lysozyme peptides detected in the LC-ECD-MS/MS analyses had sequence tags of ≥6 amino acids compared with 40.8% of those detected in the LC-CID-MS/MS analyses. The results for the remaining proteins were: cytochrome c 62.2% (ECD) and 18.9% (CID); alcohol dehydrogenase 18% (ECD) and 27.2% (CID); BSA 35.6% (ECD) and 22.2% (CID); apo-transferrin 53.3% (ECD) and 22.9% (CID); and β-galactosidase 0% (ECD) and 11.0% (CID). These results suggest that LC-ECD-MS/MS generally produces longer and therefore more reliable sequence tags than LC-CID-MS/MS. When all six proteins are considered, the ECD analyses resulted in 38.5% coverage, whereas the CID analyses gave 23.8% coverage.

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