Combination of nozzle-skimmer fragmentation and partial acid hydrolysis in electrospray ionization time-of-flight mass spectrometry of synthetic peptides.
For reliable confirmation of peptide structures, nozzle-skimmer collisionally induced dissociation was found in many cases to be insufficient, and partial acid hydrolysis was employed as a complementary technique. The utility of combining these fragmentation methods is demonstrated in two examples where the complete sequences of two synthetic peptides (peptide I, MW 2290 and peptide II, MW 1482) were unambiguously determined. In a third example, three different valine deletions in a 2 kDa synthetic peptide were identified and their positions unambiguously established. A home-built electrospray ionization time-of-flight mass spectrometer with orthogonal extraction was used for these analyses. The performance of this instrument with a resolving power of up to 7500, a mass accuracy of < or = 10 ppm, and a detection limit of 1 fmol was shown to be well suited for such studies. As a substitution to conventional external calibration, a more convenient and equally accurate internal 3-point calibration is proposed, based on the low mass ions that are present in almost all peptide spectra.
Håkansson
K
K
Angström Laboratory, Division of Ion Physics, Uppsala University, Sweden.
Zubarev
R
R
Håkansson
P
P
D000595:Amino Acid Sequence; D000818:Animals; D002138:Calibration; D002417:Cattle; D006868:Hydrolysis; D007328:Insulin / Q000032:analysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000138:chemical synthesis / Q000737:chemistry*; D013373:Substance P / Q000032:analysis
D007328:Insulin; D010455:Peptides; D013373:Substance P