Influence of different proteomic protocols on degree of high-coverage identification of nonspecific lipid transfer protein 1 modified during malting
Language English Country Germany Media print
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Chymotrypsin chemistry MeSH
- Mass Spectrometry MeSH
- Hordeum MeSH
- Edible Grain chemistry MeSH
- Molecular Sequence Data MeSH
- Fatty Acid-Binding Proteins MeSH
- Proteomics methods MeSH
- Plant Proteins chemistry MeSH
- Amino Acid Sequence MeSH
- Carrier Proteins chemistry MeSH
- Trypsin chemistry MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Chymotrypsin MeSH
- Ltp1 protein, barley MeSH Browser
- Fatty Acid-Binding Proteins MeSH
- Plant Proteins MeSH
- Carrier Proteins MeSH
- Trypsin MeSH
Both top-down (combining protein separation with MS analysis of intact proteins) and bottom-up (MS analysis of digested proteins) proteomic approaches were used for detailed characterization of nonspecific lipid transfer protein from barley malt. The aim was obtaining high-coverage of the primary structure of the proteins and the determination of PTMs such as lipid adduction and glycation. Here we present an influence of 15 proteomic protocols (differing in applied separation technique, enzyme and digestion procedure) on the extent of the coverage of the protein primary structure. The most successful protocols were in-gel digestion with trypsin of alkylated protein and in-solution digestions with trypsin or trypsin/chymotrypsin mixture of the nonalkylated protein. Totally, full sequence coverage based on the PMF and 85% sequence coverage based on the peptide fragmentation including PTMs was obtained.
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