TMC dataset

Published: 9 November 2021| Version 1 | DOI: 10.17632/szh7w479gt.1
Contributor:
Jieyu Wu

Description

To examine the neuronal protein profiles of C. elegans at different ages, we took advantage of the recently developed TurboID proximity labeling system. TurboID is an engineered biotin ligase that can label proximal proteins with biotin on nucleophilic residues such as lysine. In previous studies, proximity labeling is commonly used to identify protein-protein interactions by attaching TurboID to a protein of interest. To examine the proteomic profile of neurons in vivo, we generated a transgene that expresses free TurboID in the C. elegans nervous system. Presumably, free TurboID can evenly distribute in neurons and randomly label all proteins with biotin. Neuronal proteins can then be enriched with streptavidin beads. To test this hypothesis, we first examined the distribution of free TurboID in neurons and confirmed that TurboID was evenly distributed in all regions including the soma, axons and dendrites. We decided to collect animals during days 1, 3, 6, 9 and 12 of adulthood. After streptavidin bead enrichment, we found that the labeling efficiency was similar at different ages, and all samples were then subjected to quantitative proteomics

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Steps to reproduce

1. Biotin labeling with TurboID in C. elegans. 2. Biotinylated protein enrichment with streptavidin-coated beads 3. Quantitative LC-MS/MS Analysis

Institutions

Duke University

Categories

Neural Aging, Neuron

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