The Mass Spectral Fragmentation of Peptides

When extracting sequence information from mass spectra, not only is the m/z value at which the ions occur of importance since these provide an indication of the amino acid composition of the peptide giving rise to the ion, but so is the mass difference between adjacent ions. This indicates the particular amino acid residue that has been lost and thus provides the sequence information required. The mass differences arising from each of the amino acids are shown in Table 5.6. 

4 Confirmation of Amino Acid Sequence Using the Analysis of LC-MS Data from an Enzyme Digest of a Protein 

The sequence of a globular protein was confirmed by a combination of enzymatic digestion and HPLC with both Fourier-transform infrared spectroscopy (LC-FTIR spectroscopy) and mass spectrometry [11], 

High Performance Liquid Chromatography-Mass Spectrometry 

Protein and Peptide Analysis by Mass Spectrometry, Methods in Molecular Biology, Vol. 61, 1996. 

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Figure 5.12 Structures and nomenclature of the ions formed in the mass spectral fragmentation of peptides which involve scission of the polypeptide backbone. From Chapman, J. R. (Ed.), Protein and Peptide Analysis by Mass Spectrometry, Methods in Molecular Biology, Vol. 61, 1996. Reproduced by permission of Humana Press, Inc.

Similar techniques were used for determining the structures of unknown peptides and proteins but in these cases the individual peptides were isolated and the primary sequence determined. The mass spectral fragmentation of peptides occurs principally at the peptide linkages and can give rise to six fragment ion types. A nomenclature for this fragmentation (see... 

There are several peptide alkaloids which do not fit the above described classification, namely, pandamine (56), pandaminine (57), hymenocardine (58), and lasiodine-A (59). The determination of their structures has already been referred to (3, 4) and is briefly outlined here. It was largely consequent upon chemical degradation in part because the mass spectral fragmentation patterns had not been worked out and in part because such analyses of the alkaloids or their derivatives do not, or only partly, lend themselves to the study of such fragmentation schemes as are described in Section VII. 

Fig. 7. The mechanism for formation of b andy ions during fragmentation of protonated peptides and protein. The mass spectral pattern of b and y ions are used to determine the amino acid sequence...

Schopfer, L.M., Champion, M.M., Tamblyn, N., Thompson, C.M., Lockridge, O. (2005). Characteristic mass spectral fragments of the organophosphorus agent FP-biotin and FP-biotinylated peptides from trypsin and bovine albumin (Tyr410). Anal. Biochem. 345 122-32. 

Mass spectral fragmentation patterns of alkyl and phenyl hydantoins have been investigated by means of labeling techniques (28—30), and similar studies have also been carried out for thiohydantoins (31,32). In all cases, breakdown of the hydantoin ring occurs by a-ftssion at C-4 with concomitant loss of carbon monoxide and an isocyanate molecule. In the case of aryl derivatives, the ease of formation of Ar—NCO is related to the electronic properties of the aryl ring substituents (33). Mass spectrometry has been used for identification of the phenylthiohydantoin derivatives formed from amino acids during peptide sequence determination by the Edman method (34). 

Adding a second-stage mass spectral fragmentation by employing MS-MS (cf. Section 9.2.3.4) enables one to obtain data on the actual sequence of amino acids of an unknown peptide fragment. Figure 13.25 displays the collisionally induced fragmentation pattern of a molecular ion of the... 

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