Kyushu University develops new peptide analysis method: "Accurate even for short sequences"

In conventional peptidomics analysis, the mainstream method has been to identify peptides by matching the information obtained from mass spectrometry (MS) against known protein sequence databases. Consequently, it was difficult to accurately analyze unregistered peptides or short peptides of seven residues or fewer that contain limited information.

A research group including Assistant Professor Yumiko Toyama of the Research and Development Center for Five-Sense Devices and Associate Professor Mitsuru Tanaka of the Faculty of Agriculture at Graduate School of Kyushu University has applied a coumarin derivatization MS method to MS/MS analysis. Through this, they established a novel peptidomics technology that directly decodes the sequences of low-molecular-weight peptides from mass spectrometry data without relying on existing sequence databases. The findings were published in Analytical Chemistry.

To accurately decode short peptide sequences, the team focused on a method of attaching a tag featuring a coumarin structure to the terminus of the peptide. Attaching this tag increases the sequence information obtained via mass spectrometry, enabling a more accurate analysis of short peptides that were previously difficult to read.

Specifically, performing MS/MS analysis on the peptides observed during mass spectrometry detects fragment ions (b-ions) cleaved at the peptide bonds one residue at a time, starting from the coumarin tag modified at the N-terminus. Based on the differences in molecular weight, the type of bound amino acid can be determined one by one from the N-terminal side, allowing the peptide sequence to be identified.

When comparing the methods using 132 types of standard peptides, the conventional method correctly identified only 42 out of 86 types of dipeptides and 25 out of 46 types of oligopeptides. In contrast, the new method successfully identified all 132 types of peptides correctly. Furthermore, when analyzing casein peptone, a food-derived sample, the new method identified a larger number of peptides than the conventional approach.

This method makes it possible to determine the sequences of short peptides, which have previously been overlooked, with high precision and comprehensiveness, enabling the exploration of unknown short-chain peptides present in food and biological samples. The results of this study are expected to be useful for evaluating the functionality of foods, discovering bioactive peptides, and searching for disease-related peptides.

Tanaka said, "As the saying goes, 'Measurement is the mother of science,' and outstanding analytical methods form the very foundation that supports scientific progress. The greatest joy for us as analytical chemists is to deliver tools to the world that can fully extract the information hidden within complex samples. Nothing would bring me greater joy than to see this method reach the hands of as many researchers as possible and help open up new scientific frontiers. I will continue to pursue research that makes me feel truly fulfilled as a chemical analyst."

Journal Information
Publication: Analytical Chemistry
Title: N-Terminal Coumarin Derivatization-Aided De Novo Peptide Sequencing and Its Application to Peptidomics Using LC-Trapped Ion Mobility Spectrometry-qTOF/MS
DOI: 10.1021/acs.analchem.6c01542