Identification of Peptides of Small Extracellular Vesicles from Bone Marrow-Derived Macrophages

Jiale Cheng; Jiong Zhu; Yujiao Liu; Chen Yang; Yuehui Zhang; Yuchen Liu; Chaozhi Jin; Jian Wang

doi:10.3791/65521

Journal

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Biochemistry

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Identification of Peptides of Small Extracellular Vesicles from Bone Marrow-Derived Macrophages

JoVE Journal
Biochemistry

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JoVE Journal Biochemistry

Identification of Peptides of Small Extracellular Vesicles from Bone Marrow-Derived Macrophages

1,914 Views

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08:34 min

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June 30, 2023

DOI:

10.3791/65521-v

DOI:

10.3791/65521-v

•

08:34 min

June 30, 2023

•

21 Views

Jiale Cheng^1,2, Jiong Zhu^1,2, Yujiao Liu^2,3, Chen Yang^2,3, Yuehui Zhang², Yuchen Liu², Chaozhi Jin², Jian Wang^1,2

¹School of Basic Medical Sciences,Anhui Medical University, ²State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing),Beijing Institute of Lifeomics, ³School of Life Sciences,Hebei University

Transcript

We explore the functional roles of extracellular vesicles in innate immunity. We have developed a protocol to isolate small extracellular vesicles, or sEVs, via differential ultracentrifugation and identify the peptidome by LC-MS/MS, revealing the key biological functions of the peptides in sEVs. The extracellular vesicles have been shown to mediate intercellular communication by transporting cargo to recipient cells, such as proteins, lipids, and nucleic acids.

By releasing antimicrobial components, they act the first line of defense against invading microbes in innate immunity. While differential ultracentrifugation yields highly pure small extracellular sEVs, it can be time-consuming and often results in low yields. In our study, these are major challenges for the stable identification of low-abundance peptides in the sEVs.

Summary

This protocol describes a procedure to isolate small extracellular vesicles from macrophages by differential ultracentrifugation and extract the peptidome for identification by mass spectrometry.