De novo analysis of protein N-terminal sequence utilizing MALDI signal enhancing derivatization with Br signature
| Autor: | Seung Bum Park, Jong-Seo Kim, Yongju Kim, Hie Joon Kim, Jin Su Song |
|---|---|
| Rok vydání: | 2011 |
| Předmět: |
Phosphopeptides
Signal peptide Stereochemistry Molecular Sequence Data Peptide Peptide Mapping Biochemistry Analytical Chemistry Hemoglobins chemistry.chemical_compound Peptide mass fingerprinting Sequence Analysis Protein Imidoesters Chymotrypsin Amino Acid Sequence Phosphorylation chemistry.chemical_classification Chromatography Myoglobin Phosphopeptide Lysine Proteins Genome project Bromine N-terminus chemistry Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Mass spectrum |
| Zdroj: | Analytical and Bioanalytical Chemistry. 402:1911-1919 |
| ISSN: | 1618-2650 1618-2642 |
| Popis: | De novo analysis of protein N-terminal sequence is important for identification of N-terminal proteolytic processing such as N-terminal methionine or signal peptide removal, or for the genome annotation of uncharacterized proteins. We introduce a de novo sequencing method of protein N terminus utilizing matrix-assisted laser desorption/ionization (MALDI) signal enhancing picolinamidination with bromine isotopic tag incorporated to the N terminus. The doublet signature of bromine in the tandem mass (MS/MS) spectrum distinguished N-terminal ion series from C-terminal ion series, facilitating de novo N-terminal sequencing of protein. The dual advantage of MALDI signal enhancement by the basic picolinamidine and b-ion selection aided by Br signature is demonstrated using a variety of peptides. The N-terminal sequences of myoglobin and hemoglobin as model proteins were determined by incorporating the Br tag to the N terminus of the proteins and obtaining a series of b-ions with Br signature by MS/MS analysis after chymotryptic digestion of the tagged proteins. The N-terminal peptide was selected for MS/MS analysis from the chymotryptic digest based on the Br signature in the mass spectrum. Identification of phosphorylation site as well as N-terminal sequencing of a phosphopeptide was straightforward. |
| Databáze: | OpenAIRE |
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