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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 containing sequence tags of ≥6 consecutive amino acids (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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fig3: Protein sequence coverage obtained for the six proteins containing sequence tags of ≥6 consecutive amino acids (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: The overall protein sequence coverage with six or more consecutive cleavages, averaged over the three replicate injections, can be seen in Figure 3. This shows that for four of the six proteins, ECD provided greater sequence coverage of the proteins when peptides contain sequence tags ≥6 amino acids long. The average sequence coverages obtained using these criteria were: lysozyme 12.4% (ECD) and 10.8% (CID); cytochrome c 20.4% (ECD) and 6.7% (CID); alcohol dehydrogenase 1.3% (ECD) and 5.8% (CID); BSA 11.85 (ECD) and 4.3% (CID); apo-transferrin 15.7% (ECD) and 4.9% (CID); and β-galactosidase 0% (ECD) and 1.6% (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 containing sequence tags of ≥6 consecutive amino acids (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

fig3: Protein sequence coverage obtained for the six proteins containing sequence tags of ≥6 consecutive amino acids (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: The overall protein sequence coverage with six or more consecutive cleavages, averaged over the three replicate injections, can be seen in Figure 3. This shows that for four of the six proteins, ECD provided greater sequence coverage of the proteins when peptides contain sequence tags ≥6 amino acids long. The average sequence coverages obtained using these criteria were: lysozyme 12.4% (ECD) and 10.8% (CID); cytochrome c 20.4% (ECD) and 6.7% (CID); alcohol dehydrogenase 1.3% (ECD) and 5.8% (CID); BSA 11.85 (ECD) and 4.3% (CID); apo-transferrin 15.7% (ECD) and 4.9% (CID); and β-galactosidase 0% (ECD) and 1.6% (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