Yoshio Kodera (Sagamihara / JP), Yusei Okuda (Sagamihara / JP), Yuzuru Nakagawa (Sagamihara / JP), Takashi Matsui (Sagamihara / JP), Makoto Itakura (Sagamihara / JP)
Bioactive peptides secreted by tissues circulate in the blood and perform various functions. Therefore, bioactive peptides are used as drugs and biomarkers. To identify novel bioactive peptides, we developed the Differential Solubilization (DS) method, a highly efficient peptide extraction method for plasma. As a result, we were able to identify more than 10,000 peptides. Howverer, most of the identified peptides were fragments of major proteins in plasma. It was considered necessary to identify peptides in plasma based on tissue-derived peptides for the discovery of novel bioactive peptides. Therefore, we started peptidome analysis in mouse tissues with the aim of proposing novel bioactive peptides from tissue-derived peptides in plasma.
A mouse tissue was frozen immediately after dissection. Peptides were extracted from a frozen tissue using the modified differential solubilization (DS) method1,2 optimized for tissue and analyzed using Q-Exactive equipped with an EASY-nLC 1000 system (Thermo Fisher Scientific). LC-MS/MS data were searched using PEAKS Studio (Bioinformatics Solutions Inc.).
For the hypothalamic tissue, LC-MS/MS analysis resulted in the identification of 1,535 peptides derived from 297 proteins. Approximately 45% of the identified peptides (690 of 1,535 peptides) belonged to prohormone precursor protein groups. Within the prohormone precursor protein group, 35 bioactive peptides listed in the database, including substance P, neurokinin A, neuropeptide K, neuropeptide Y, lipotropin γ and α-MSH, were identified. Furthermore, as for the neuropeptides secreted from the hypothalamus, Orexin B, β-endorphin and three opioids α-neoendorphin, dynorphin A and dynorphin B were detected.
In the presentation, the results of peptidome analysis of the liver, kidney and brain tissue other than the hypothalamus will be presented and discussed in comparison with the results of previous peptide analyses.
1. Kawashima Y., et al. 2010 J. Proteome Research. 9:1694-705
2. Nakagawa Y., et al. 2021 Biochem Biophys Res Commun. 548:155-160