Introduction
The analysis process of recombinant protein expression and purification products requires confirmation of the full
length of the protein, especially the development and process establishment of recombinant protein products or
antibody agents. Determining the complete sequence of the protein is helpful to analyze the structure of the protein
and reveal the biological function of the protein. In traditional "bottom-up" proteomic strategy, database
searching is used to identify sequences of peptides and proteins analyzed by liquid chromatography-tandem mass
spectrometry (LC-MS/MS). Protein sequences absent from the database cannot be identified, and even if present in the
database, complete sequence coverage is rarely acheved even for the most abundant proteins in the sample.
Fig.
1. Full-length protein sequencing based on continuous digestion using non-specific proteases. (Yang C, et
al., 2021)
Our Services
Creative Proteomics has developed a full-length protein sequencing method based on continuous
digestion using various non-specific proteases by utilizing the existing high-resolution mass spectrometry platform,
which chould increase the accuracy and coverage of protein sequencing. A continuous digestion device was
constructed, and a variety of non-specific proteases were used to continuously digest the protein. Taking advantage
of the non-specific cleavage sites of non-specific proteases, the complementarity of peptides produced at different
time and by different kinds of proteases, the type and overlapping degree of digested peptides were improved. The
sequence coverage of peptides after continuous digestion by each protease can reach 100%. This analysis can be used
to confirm whether the recombinant protein is fully expressed and to detect whether the recombinant protein
expression process is broken.
Creative Proteomics has developed the following a customized sequencing service process for protein
full-length sequencing analysis services.
Ⅰ. Predicted sequence of a protein based on the gene are analyzed to selecte different enzymes resulting in
complement digestive sites.
Ⅱ. Protein are digested separately by selected enzymes.
Ⅲ. The previous step digested pepiteds are analyzed by HPLC-MS/MS.
Ⅳ. MS data are analyzed and pieced together for full-length protein sequence.
Sample requirements
- Gel samples and solution samples could be used for the above protein full-length sequencing.
- Amount: 10-20 μg.
- Purity of protein samples should be as high as possible.
Our Technical Advantages
- Peptide coverage up to 100%.
- High sensitivity.
- A formal report, reporting the experimental process, protein full sequence information, and secondary map
support.
Applications
- Determination of Full-Length Sequence of Biologics (Proteins/Peptides/Antibodies/Vaccines)
- Analysis of PTMs
- Verification of the Correct Translation of Recombinant Proteins
Please feel free to contact us If you would like to discuss the best way to
determine the protein full-length sequencing of your protein. At Creative Proteomics, we not only
provide professional protein full-length sequencing service projects for global customers, our scientists can also
provide customized services according to your specific requirements, and describe them in detailed project reports,
we look forward to with your cooperation.
References
- Yang C, Shan Y C, Zhang W J, et al. (2021) Full-length Protein Sequencing Based on Continuous Digestion
Using Non-specific Proteases. Acta Chimica Sinica. 79(5): 663-669.
