PMF for MALDI-TOF

Pechar

A process of reduction and carbamidomethylation from proteins in solution, or in gel, followed by enzymatic digestion with trypsin (an enzyme that specifically cleaves lysines and arginines, if not followed by a proline), is carried out to obtain their peptides.

The tryptic digest is analysed using the MALDI-TOF technique from which the m/z ratio of the peptides generated in the digestion is obtained, which will determine the peptide fingerprint of that protein (PMF). These experimental masses will be compared with the theoretical masses in the databases and their correlation will lead to an identification, using a search engine (PEAKS, MASCOT).

When the identification by PMF analysis fails, an analysis by MALDI- TOF/ TOF technique is performed. In this way, protein identification will be obtained by fragmentation spectra of some of the peptide masses obtained and their correlation with existing databases.

Low molecular weight proteins (<20 KDa) can be difficult to identify, due to the low number of peptides obtained from their digestion.