日本質量分析学会 第66回質量分析総合討論会

プログラム

ポスター発表

第2日 5月16日(水)  ポスター会場

Identification of protein targets of a bioactive small molecule with DARTS combined with LC-MS/MS proteome analysis

(1Yonsei University2Severance Biomedical Science3Korea Basic Science)
oLee, Dongjin1Hwang, Hui-yun1Lee, Myung-shik2Lim, Hyejin2Kim, Jin Young3Yoo, Jong Shin3Kwon, Ho Jeong1

Identification of protein target of bioactive small molecules provides the basis for the understanding of its mode of actions. We have developed a new label-free target protein identification method by combination of DARTS (Drug Affinity Responsive Target Stability) with LC-MS/MS. Recently, we found out a new small molecule (MSL) enhancing autophagy activity through TFEB (transcription factor EB) translocation and identified Calcineurin A (CaN)as a protein target of MSL through DARTS assay. We further validated the specific interaction of MSL with CaN by comparing the binding activities of inactive derivatives of MSL to CaN. Through LC-MS/MS proteome analysis of the entire proteome treated with MSL with pronase, 20 proteins exhibited resistant to protease upon MSL treatment. Molecular and cellular investigations revealed that MSL could bind to other phosphatases like CaN for TFEB translocation into nucleus leading to activation of autophagy. These results provide the role of phosphatases in autophagy induction via TFEB regulation and combination of DARTS and LC-MS/MS method could be an efficient platform to identify protein targets of unmodified small molecules.