Practical Protein Chemistry - A. Darbre 1989

Peptide mapping of proteins
Peptide mapping — types of structures analyzed

Below are the most likely scenarios encountered when performing comparative analysis using two-protein mapping.

1) Identical structures: Amino acid sequences and post-translational modifications (if any) are completely homologous.

2) The amino acid sequences are homologous, but differences exist in the modification of side chains or terminal residues (Acetylation, glycosylation, phosphorylation, etc.). See the review on protein modification [72].

3) The amino acid sequences are identical, but one protein is shorter than the other at the N- or C-terminus, or at both ends.

4) Part of the sequence in both Proteins matches completely, while another part exhibits total or partial differences.

5) Significant Regions of the sequence are homologous, but insertions or deletions are irregularly distributed along the chain.

6) Complete structural mismatch.

Naturally, various combinations of these basic types may occur.

The simplest of all considered scenarios are the first and the last, which yield entirely identical or completely non-matching maps, respectively. It should be noted that the reliability of comparative analysis increases with the number of components contained in the test sample, as this provides more data for investigation.

Proteins with homologous amino acid sequences that differ in post-translational modifications (type 2) yield similar maps, with the sole exception of those fragments whose mobility is altered by the corresponding modification. A similar pattern (partial chromatogram overlap) is observed in scenario 4. Detecting structural differences using one-dimensional gel Electrophoresis, which analyzes large Polypeptides, is less likely than using peptide maps, because at a low molecular weight, changes in Structure/106.html">Hydrophobicity or charge have a significantly greater impact on peptide behavior.

If one of the two proteins is a fragment of the other (type 3), the peptide map of the shortened sample will differ only by the absence of one or more fragments. Furthermore, one fragment in each map will not be shared—specifically, the one adjacent to or resulting from the Cleavage site. If a protein is shortened relative to the other at both the N- and C-termini, there will be two such non-matching fragments in each map.

Perhaps the most complex situation arises when comparing peptide maps of proteins that have similar amino acid sequences but contain irregular substitutions, insertions, or deletions (type 5) resulting from Gene Duplication events. Naturally, the peptide maps of two such molecules will share more similarity the fewer such changes have occurred. However, even significant Homology of this type is not always obvious. For instance, the amino acid sequences of Acetylcholine Receptor subunits exhibit ~40% homology [51] and yield distinct peptide maps [32] (Fig. 7.1). It should be borne in mind that in such cases the method's capabilities are limited, and careful Interpretation of Results is essential to avoid potential errors.