HI. I am trying to inhibit ubiquitination in Human Brain Endothelial Cells In Vitro using PYR-41 ubiquitin ligase inhibitor. But it is not inhibiting ubiquitination as lower concentration and at higher concentration it is too toxic to my cells.
Is there any suggestion to use PYR-41 or any other good ubiquitin ligase inhibitor.
Dear Smita,
PYR-41 is an inhibitor of the ubiquitin activating enzyme, and will for this reason inhibit the activity of all E2's and E3's as well. For inhibiting the E1, I would recommend that you use MLN7243, which is a substrate assisted ubiquitin E1 inhibitor, and very potent both in vitro and in cell culture. Please see this for reference: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3220466/
The inhibitor is available here:
http://www.activebiochem.com/Product/MLN-7243.html
Best of luck with your experiments,
Roland.
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