Structures of peptide fragments

No tandem MS experiment can be successful if the precursor ions fail to fragment (at the right time and place). The ion activation step is crucial to the experiment and ultimately defines what types of products result. Hence, the ion activation method that is appropriate for a specific application depends on the MS instrument configuration as well as on the analyzed compounds and the structural information that is wanted. Various, more or less complementary, ion activation methods have been developed during the history of tandem MS. Below we give brief descriptions of several of these approaches. A more detailed description of peptide fragmentation mles and nomenclature is provided in Chapter 2. An excellent review of ion activation methods for tandem mass spectrometry is written by Sleno and Volmer, see Reference 12, and for a more detailed review on slow heating methods in tandem MS, see Reference 13. 

Structural similarities of peptidic fragments contained in primary protein structures or structural relationships found for some protein pairs are in some cases of such a character as to be almost immediately obvious (cf., e.g., the heme protein from Chromatium). However, the question whether all similarities considered are real should be established by a rigorous application of the calculus of probability. Two attempts in this direction were made recently the first one, by Williams et al. (1961), is experimental in nature while the second reported by orm and Knichal (1962) represents a theoretical approach. 

Figure 2 Nomenclature of peptide fragmentation. The possible product ion series that arise by cleavages along the peptide backbone are a-, b-, and c-series (N-terminal) and x-, y-, and z-series (C-terminal). (The designation +2H denotes addition of two hydrogens that are transferred onto the structures depicted in the figure to form the corresponding singly charged y- or c- product ions (21)). Under low-energy CID, y- and b-ions usually predominate. The mass differences between adjacent ions of the same series can be used to deduce portions of the peptide sequence.

II. Predictions of PrP Structures and Studies of Peptide Fragments to Test the Models... 

Fig. 3. Molecular dynamics simulations of peptide fragments corresponding to the predicted secondary structure in Fig. 2. H2.5 was discovered while trying to model the missing residues from the PrP prediction. All simulations began with the peptide in the a-helical conformation, and the average percentage of helix over the 2 ns simulation is given.

In order to determine the sequence of amino acids in a peptide, initially it was subjected to partial hydrolysis. Thereby fragments were formed, smaller peptides the sequence of which could be more easily established. After micropreparative separation, e.g. by paper chromatography, the fragments were hydrolyzed and analyzed for amino acid composition both before and after treatment with nitrous acid. The amino add missing in the treated, desaminated, sample, was its N-terminal residue. The sequence of the whole peptide was then reconstructed through the appropriate combination of the structurally identified peptide fragments. A typical example, the eluddation of the structure of gramiddin S, is described on p. 205. 

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