Redirecting the specificity of tripartite motif containing-21 scaffolds using a novel discovery and design approach.
E3 ubiquitin ligase
TRIM21
bioPROTAC
directed evolution
lymphoma
protein degradation
scaffold protein
targeted protein degradation
Journal
The Journal of biological chemistry
ISSN: 1083-351X
Titre abrégé: J Biol Chem
Pays: United States
ID NLM: 2985121R
Informations de publication
Date de publication:
21 Oct 2023
21 Oct 2023
Historique:
received:
05
04
2023
revised:
30
09
2023
accepted:
16
10
2023
pubmed:
23
10
2023
medline:
23
10
2023
entrez:
22
10
2023
Statut:
aheadofprint
Résumé
Hijacking the ubiquitin proteasome system to elicit targeted protein degradation (TPD) has emerged as a promising therapeutic strategy to target and destroy intracellular proteins at the post-translational level. Small molecule-based TPD approaches, such as proteolysis-targeting chimeras (PROTACs) and molecular glues, have shown potential, with several agents currently in clinical trials. Biological PROTACs (bioPROTACs), which are engineered fusion proteins comprised of a target-binding domain and an E3 ubiquitin ligase, have emerged as a complementary approach for TPD. Here, we describe a new method for the evolution and design of bioPROTACs. Specifically, engineered binding scaffolds based on the third fibronectin type III domain of human tenascin-C (Tn3) were installed into the E3 ligase tripartite motif containing-21 (TRIM21) to redirect its degradation specificity. This was achieved via selection of naïve yeast-displayed Tn3 libraries against two different oncogenic proteins associated with B-cell lymphomas, mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) and embryonic ectoderm development protein (EED), and replacing the native substrate-binding domain of TRIM21 with our evolved Tn3 domains. The resulting TRIM21-Tn3 fusion proteins retained the binding properties of the Tn3 as well as the E3 ligase activity of TRIM21. Moreover, we demonstrated that TRIM21-Tn3 fusion proteins efficiently degraded their respective target proteins through the ubiquitin proteasome system in cellular models. We explored the effects of binding domain avidity and E3 ligase utilization to gain insight into the requirements for effective bioPROTAC design. Overall, this study presents a versatile engineering approach that could be used to design and engineer TRIM21-based bioPROTACs against therapeutic targets.
Identifiants
pubmed: 37866632
pii: S0021-9258(23)02409-2
doi: 10.1016/j.jbc.2023.105381
pmc: PMC10694607
pii:
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
105381Informations de copyright
Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Conflict of interest J.H. and R.G. are employees and hold shares in AstraZeneca. All the authors declare that they have no conflicts of interest with the contents of this article.
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