Document Type

Article

Department/Program

Biology

Journal Title

Journal of Visualized Experiments

Pub Date

4-27-2019

Publisher

JOVE

Volume

146

Journal Article URL

https://www.jove.com/video/59576/localization-sumo-modified-proteins-using-fluorescent-sumo-trapping

Abstract

Here we are presenting a novel method to study the sumoylation of proteins and their sub-cellular localization in mammalian cells and nematode oocytes. This method utilizes a recombinant modified SUMO-trapping protein fragment, kmUTAG, derived from the Ulp1 SUMO protease of the stress-tolerant budding yeast Kluyveromyces marxianus. We have adapted the properties of the kmUTAG for the purpose of studying sumoylation in a variety of model systems without the use of antibodies. For the study of SUMO, KmUTAG has several advantages when compared to antibody-based approaches. This stress-tolerant SUMO-trapping reagent is produced recombinantly, it recognizes native SUMO isoforms from many species, and unlike commercially available antibodies it shows reduced affinity for free, unconjugated SUMO. Representative results shown here include the localization of SUMO conjugates in mammalian tissue culture cells and nematode oocytes.

DOI

10.3791/59576

Available for download on Tuesday, April 27, 2021

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