C-terminal ladder sequencing by an approach combining chemical degradation with analysis by matrix-assisted-laser-desorption ionization mass spectrometry.
A chemical procedure for the degradation of peptides and analysis by matrix-assisted-laser-desorption ionization mass spectrometry (MALDI-MS) was used for C-terminal sequence determination. The method allowed us to determine up to eight amino acid residues in the lower picomole range by mass analysis without any repetitions of the degradation nor any extraction or purification of the truncated peptide chains. The C-terminal degradation of all 20 common amino acid residues was proved by applying this method. Extended C-terminal sequence information from enzymatic digests using carboxypeptidase P was obtained by combining the enzymatic with the chemical mass spectrometric approach. Furthermore the amino acids lysine and glutamine, with the same masses, were distinguishable due to the formation of acetyl-lysine in the chemical process.
Thiede
B
B
Max-Delbrueck-Center of Molecular Medicine, Protein Chemistry Research Group, Berlin-Buch, Germany.
Salnikow
J
J
Wittmann-Liebold
B
B
D000595:Amino Acid Sequence; D000818:Animals; D002268:Carboxypeptidases; D005069:Evaluation Studies as Topic; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification; D017421:Sequence Analysis / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013379:Substrate Specificity
D010446:Peptide Fragments; D010455:Peptides; C025427:carboxypeptidase P; D002268:Carboxypeptidases