Monday, February 27, 2006

post-translational modifications

General Approach
Most protein samples exhibit some degree of modification.
There are the "natural" post translational modifications, such as phosphorylation and glycosylation. There are the accidental modifications which are artefacts of sample handling, such as oxidation. Finally, there are the modifications deliberately introduced during sample work-up, such as cysteine derivatisation. In most cases, it is only the deliberate modifications which are known about for certain at the time of doing a search.
It might be assumed that the search software could allow for those modifications which are described in sequence entry annotations. However, writing code to parse these sequence annotations would be a major task. Indeed, many post-translational modifications are not specified in a way which can be readily translated into specific mass differences. For example, noting that a residue is an actual or potential glycosylation site is not much help. Even a simple modification, such as phosphorylation, is rarely quantitative, so that it would be necessary to include mass values for all permutations of occupied and unoccupied sites.
And, of course, protein sequences derived translated from nucleotide sequences contain no information on post translational modifications.
The solution adopted here is to allow modifications to be specified in two different ways: fixed modifications and variable modifications.
Fixed modifications are applied universally, to every instance of the specified residue or terminus. There is no computational overhead associated with a fixed modification, it is simply equivalent to using a different mass for the modified residue or terminus. For example, selecting Carboxymethyl (C) means that all calculations will use 161 Da as the mass of cysteine.
Variable modifications are those which may or may not be present. Mascot tests all possible arrangements of variable modifications to find the best match. For example, if Oxidation (M) is selected, and a peptide contains 3 methionines, Mascot will test for a match with the experimental data for that peptide containing 0, 1, 2, or 3 oxidised methionine residues. This greatly increases the complexity of a search, resulting in longer search times and reduced specificity, so variable modifications should be used sparingly.
The list of modifications used by Mascot is taken directly from the Unimod database. For further details of individual modifications, please refer to Unimod. Note that Unimod is an community supported resource. If you want to add a new modification to Unimod, you can do so, and you then become the curator of the new record. The Mascot modifications list is updated from Unimod each weekend.
Only selected modifications are displayed by default in the Mascot search form. If you want to see the complete list, you must go to the search form defaults page and tick the checkbox for 'Show all mods.'.
Other lists of modifications
DeltaMass is a comprehensive list of modifications, sorted by mass.
RESID database contains detailed descriptions of many post-translational modifications.


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