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.
another two more questions :1 , Does all the E3s have the ability to polyubiquitylate itself ?? 2, we assume the E3 can catalyse k63-linked polyub. How can i choose E2 except the E2 kit ?
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