Creative Proteomics provides N-terminal sequence analysis by both Edman and Mass spectrometry of therapeutic proteins, monoclonal antibodies and protein vaccines.
N-terminal Edman degradation:
In protein N-terminal sequence analysis, proteins are first modified with phenylisothiocyanate (PITC). Subsequently, the derivatized terminal amino acid is removed by acid cleavage in a form of phenylthiohydantoin (PTH) derivative and a new α-amino group on the next amino acid is now available for the next round of reaction with PITC. Protein sequence is therefore being analyzed in through a serious reaction of PITC addition, and cleavage of one PTH at each time for analysis. Suppose the reactions are fully efficient, this method can sequence the whole amino acid from the N-terminal end. Though less efficient as compared with mass spectrometer, N-terminal Edman sequencing still has advantages for protein analysis that cannot readily be obtained by other analysis methods. With current technology, it is fairly routine to obtain at least 20 to 25 residues of sequence from the N-terminus of the proteins and peptides.
Mass spectrometry:
Currently, the most commonly used technique for protein sequence analysis is mass spectrometry. Mass spectrometry is a highly efficient method for the accurate mass determination and characterization of proteins. Two mainly used mass spectrometers are electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI), respectively. Basically, proteins are pretreated with digestive enzymes to be digested into small fragments, which are analyzed by mass spectrometers. Depends on the projects need, MALDI and ESI are two used as ionization source for ionizing peptide fragments. Usually, MALDI is more frequently used when dealing with large amount of samples. Peptide sequences are then obtained by analyzing the mass spectrum of each of the fragments, which together consist of the full-length protein sequence.
Protein De Novo Sequencing
The biggest feature of the protein de novo sequencing method is that it can directly analyze the protein sequence without using any protein or DNA database information. Compared with the traditional mass spectrometry database search method, it has an incomparable advantage, that is, the de novo sequencing method can be applied to analyze the protein sequences of new species, as well as the protein sequences of species whose genomes have not been sequenced.
Application of protein sequencing service:
- Analysis of protein N-, or C-terminal signal sequence
- Protein N-, or C-terminal tags
- Post-translational modifications on N-, or C- terminal ends
- Full-length sequence detection of proteins and peptides with multiple amino acid residues (such as hundreds or more);
- Monoclonal antibody full-length sequencing
Technology platform:
- Liquid Chromatography (LC)
- High Performance Liquid Chromatography (HPLC)
- Matrix Assisted Laser Desorption Ionization Mass Spectrometry (MALDI-MS)
- Nano HPLC- 7 T solariX XR FTICR-MS (equipped with ECD)
- Nano HPLC- Orbitrap Fusion™ Lumos™ Tribrid™ MS (equipped with ETD)
Ordering Procedure:
