一种过氧标签可实现从人细胞中快速分离过氧化物酶体。
A PEROXO-Tag Enables Rapid Isolation of Peroxisomes from Human Cells.
作者信息
Ray G Jordan, Boydston Elizabeth A, Shortt Emily, Wyant Gregory A, Lourido Sebastian, Chen Walter W, Sabatini David M
机构信息
Whitehead Institute for Biomedical Research, 455 Main Street, Cambridge, MA 02142, USA; Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Department of Biology, 77 Massachusetts Avenue, Cambridge, MA 02139, USA; Broad Institute of Harvard and Massachusetts Institute of Technology, 415 Main Street, Cambridge, MA 02142, USA.
Whitehead Institute for Biomedical Research, 455 Main Street, Cambridge, MA 02142, USA.
出版信息
iScience. 2020 May 22;23(5):101109. doi: 10.1016/j.isci.2020.101109. Epub 2020 Apr 28.
Peroxisomes are metabolic organelles that perform a diverse array of critical functions in human physiology. Traditional isolation methods for peroxisomes can take more than 1 h to complete and can be laborious to implement. To address this, we have now extended our prior work on rapid organellar isolation to peroxisomes via the development of a peroxisomally localized 3XHA epitope tag ("PEROXO-Tag") and associated immunoprecipitation ("PEROXO-IP") workflow. Our PEROXO-IP workflow has excellent reproducibility, is easy to implement, and achieves highly rapid (~10 min post homogenization) and specific isolation of human peroxisomes, which we characterize here via proteomic profiling. By offering speed, specificity, reproducibility, and ease of use, the PEROXO-IP workflow should facilitate studies on the biology of peroxisomes.
过氧化物酶体是在人体生理学中执行各种关键功能的代谢细胞器。传统的过氧化物酶体分离方法可能需要1个多小时才能完成，且实施起来可能很费力。为了解决这个问题，我们现在通过开发一种过氧化物酶体定位的3XHA表位标签（“PEROXO标签”）和相关的免疫沉淀（“PEROXO-IP”）工作流程，将我们之前关于快速细胞器分离的工作扩展到过氧化物酶体。我们的PEROXO-IP工作流程具有出色的可重复性，易于实施，并且能够高度快速（匀浆后约10分钟）和特异性地分离人过氧化物酶体，我们在此通过蛋白质组学分析对其进行表征。通过提供速度、特异性、可重复性和易用性，PEROXO-IP工作流程应有助于过氧化物酶体生物学的研究。
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