De novo sequencing: From Edman to mass spectrometry
Mario Cindrić, Ruđer Bošković Institute.
Novel method for peptide amino acid sequence determination upgraded with genome fingerprint scanning (GFS) was designed to investigate gene expression of the clinical organisms at the first instance but it can be used in further development as ultimate de novo sequencing method in “bottom up” proteomics.
The method links proteomics data, consisting of determined tryptic peptides amino acid sequences obtained by negative and positive ion modes. MALDI MS/MS sequencing data read and compared to sequenced genome of the observed organism is confirmed by overlapping of the sequence data from both modes (e.g. GAAGAK read in neg. and pos. ion modes; GAAGAK b ions neg. ion mode and KAGAAG y ions pos. ion mode).
The idea that lies behind this new technology is enhanced de novo sequencing of unknown peptide amino acid sequences in negative MS/MS (enabled by derivatization that contains two negatively charged groups without observed side reactions or peptide degradation) and positive MS/MS of the same peptide used as quality control (MS/MS of derivatized ions in both cases).
