Introduction
In spite of the continuously growing sequence databases, de novo sequencing of peptides, i.e. sequencing without
assistance of a linear sequence database, is still essential in several analytical situations. For example, analyses
of protein sequence variants or their splice isoforms require de novo sequencing, as well as protein analysis from
organisms with unsequenced genomes. In addition, de novo sequencing is essential for analysis of peptides containing
non-proteinic or modified amino acids, as typically present. The basic principle of de novo sequencing is to use the
mass difference between two fragment ions to calculate the mass of amino acid residues on the peptide chain, among
which CID and ETD are the most effective techniques for de novo peptide sequencing, both of which can generate
Peptide MS/MS spectra with very high structural information, CID is best for small peptides of 1-2 kDa, while ETD
can handle larger peptides.
Fig. 1. The
process of peptide de novo sequencing by mass spectrometry. (Hao Y, et al. 2019)
Our Services
Creative Proteomics has developed a combined HCD+ETD peptide de novo sequencing method by using mass
spectrometry to obtain higher energy collisional dissociation (HCD) and electron transfer dissociation (ETD) spectra
of the same precursor. Our experienced scientists using the combination of HCD+ETD that it can collect more
complementary fragmentation information with high resolution and high-quality accuracy to greatly improve the
accuracy and speed of de novo sequencing. 86% of the top-most candidate sequences derived by our method from the HCD
+ ETD spectral pair matched the database search results with 95% accuracy, much higher than using only HCD (87%) or
only ETD spectra (57%), the average peptide sequence extraction time is only 0.018s.
In addition, we will use a variety of proteases to digest and identify the target peptides respectively. While
obtaining the fragmented peptide fragments, the splicing between the peptide fragments will complete the 100%
determination of the peptide sequence, that is, by De novo sequencing method was used for the determination.
Our Technical Advantages
Our established peptide de novo sequencing platform has been widely chosen by our cusyomer for the following unique
features.
- Several proteases commonly are used for the enzymatic cleavage and mass spectrometry of the target protein
respectively, and the determination of 100% of the protein sequence was completed by splicing between peptides
while obtaining fragmented peptide fragments.
- The combination of HCD and ETD is adopted to ensure the integrity of the fragmented peptide fragments in the
process of peptide fragmentation.
- Advanced data processing algorithms is used to parse the raw data, for < 30 amino acids are generally
deducedby the database or de novo de novo sequencing, while >30 amino acids are generally deduced by de novo
sequencing.
At Creative Proteomics, we provide complete and professional peptide de novo sequencing services for
global customers which was used for the discovery of new peptides and development of multifunctional peptides and
complexes. Our professional scientists can also design customized services according to your requirements. Please
feel free to contact us If you are interested in our services, we look
forward to with your cooperation.
References
- Seidler J, Zinn N, Boehm M E, et al. (2010) De novo sequencing of peptides by MS/MS.
Proteomics. 10(4):634-49.
- Yang H, Li YC, Zhao M Z, et al. (2019) Precision de Novo peptide sequencing using mirror proteases of
ac-lysargiNase and trypsin for large-scale proteomics. Mol Cell Proteomics.18(4):773-785.
