1. Does the ubiquitination of k48-linkage or k63-linkage share the same E2 in some case ?
2. How to choose a right E2 when identifying whether a protein can act as an E3 ligase for k63-linkage ubiquitination? Do I need to perform in vitro Ubiquitination assay to scan all the potential E2?
thx for advice !
Hello,
1. Yes the same E2 can form k48-linkages and k63 linkages. To my knowledge the different linkages depend on the E3 and substrate.
2. Abcam sells a E2-Ubiquitin Conjugation Kit with multiple human recombinant E2s if you are trying to determine a which E2s are compatible with your E3.
Human UbcH5 and Arabidopsis Ubc8 are gold standard E2 enzymes that tend to be active with the broadest array of RING-type E3 ligases. As stated above, chain topology usually depends on the E3-substrate interaction. If you are performing an in vitro reaction, keep in mind that protein concentration and buffer additives can also influence the nucleophilicity and accessibility of residues.
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