Nonradioactive quantification of autophagic protein degradation with L-azidohomoalanine labeling

@article{Wang2017NonradioactiveQO,
  title={Nonradioactive quantification of autophagic protein degradation with L-azidohomoalanine labeling},
  author={Jigang Wang and Jianbin Zhang and Yew Mun Lee and Shukie Ng and Yin Shi and Zi-Chun Hua and Qingsong Lin and H M Shen},
  journal={Nature Protocols},
  year={2017},
  volume={12},
  pages={279-288}
}
At present, several assays that use radioisotope labeling to quantify the degradation of long-lived proteins have been developed to measure autophagic flux. Here, we describe a nonradioactive pulse–chase protocol using L-azidohomoalanine (AHA) labeling to quantify long-lived protein degradation during autophagy. AHA is used as a surrogate for L-methionine, and, when added to cultured cells grown in methionine-free medium, AHA is incorporated into proteins during de novo protein synthesis. After… CONTINUE READING
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