8J0A

Robust design of effective allosteric activator UbV R4 for Rsp5 E3 ligase using the machine-learning tool ProteinMPNN

Summary for 8J0A

• Entry DOI: 10.2210/pdb8j0a/pdb
• Descriptor: Ubiquitin variant R4, SULFATE ION (3 entities in total)
• Functional Keywords: ubiquitin variant, deep-learning, biosynthetic protein
• Biological source: synthetic construct
• Total number of polymer chains: 2
• Total formula weight: 23620.53

Authors

Lin, Y.-F.,Hsieh, Y.-J.,Kao, H.-W.,Ko, T.-P.,Wu, K.-P. (deposition date: 2023-04-10, release date: 2023-08-09, Last modification date: 2023-08-30)

Primary citation

Kao, H.W.,Lu, W.L.,Ho, M.R.,Lin, Y.F.,Hsieh, Y.J.,Ko, T.P.,Danny Hsu, S.T.,Wu, K.P.
Robust Design of Effective Allosteric Activators for Rsp5 E3 Ligase Using the Machine Learning Tool ProteinMPNN.
Acs Synth Biol, 12:2310-2319, 2023

PubMed Abstract: We used the deep learning tool ProteinMPNN to redesign ubiquitin (Ub) as a specific and functionally stimulating/enhancing binder of the Rsp5 E3 ligase. We generated 20 extensively mutated─up to 37 of 76 residues─recombinant Ub variants (UbVs), named R1 to R20, displaying well-folded structures and high thermal stabilities. These UbVs can also form stable complexes with Rsp5, as predicted using AlphaFold2. Three of the UbVs bound to Rsp5 with low micromolar affinity, with R4 and R12 effectively enhancing the Rsp5 activity six folds. AlphaFold2 predicts that R4 and R12 bind to Rsp5's exosite in an identical manner to the Rsp5-Ub template, thereby allosterically activating Rsp5-Ub thioester formation. Thus, we present a virtual solution for rapidly and cost-effectively designing UbVs as functional modulators of Ub-related enzymes.

PubMed: 37556858
DOI: 10.1021/acssynbio.3c00042

Experimental method

X-RAY DIFFRACTION (3 Å)