| Protein analysis by peptide mass maps has
become an important technique for proteomic studies. A
protein from a gel spot or a separated subcellular fraction
is proteolytically digested and analyzed by LC-MS/MS.
Proteins are identified by comparison of measured peptide
fragment ions with those predicted by in silico digestion
of a protein database. This technique can account for
some modifications to the protein primary sequence, i.e.
post-translational modifications (PTMs), but the protein
must be present in the database and the modification must
be known a priori.
Locations and structures of unexpected PTMs can be
determined by further fragmentation of the modified
peptide (e.g. MS to 3 in an ion trap). They can also
be identified by analysis of existing data using enhanced
software tools for spectral interpretation, de novo
sequencing, and spectral pattern analysis. The confidence
of all of these techniques is enhanced with access to
data sets with very high mass accuracy, e.g. <2ppm.
In this report, we will demonstrate use of an ion
trap mass spectrometer (LTQ FT), coupled with protein
identification (TurboSEQUEST®), de novo sequencing
(DenovoXTM), and pattern analysis (SALSA) software to
study phosphorylation, glycosylation, and methylation
in a complex biological sample mixture. |
