Lysineless HiBiT and NanoLuc Tagging Systems as Alternative Tools Monitoring Targeted Protein Degradation DOI Open Access
Hanfeng Lin, Kristin M. Riching,

May Poh Lai

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: May 17, 2024

Target protein degradation (TPD) has emerged as a revolutionary approach in drug discovery, leveraging the cell's intrinsic machinery to selectively degrade disease-associated proteins. Proteolysis-Targeting Chimeras (PROTACs) exemplify this strategy, exploiting heterobifunctional molecules induce ubiquitination and subsequent of target The clinical advancement PROTACs underscores their potential therapeutic intervention, with numerous projects progressing through stages. However, monitoring subtle changes abundance induced by TPD demands highly sensitive assays. Nano-luciferase (nLuc) fusion proteins, or NanoBiT technology derived from it, offer robust screening platform due high sensitivity stability. Despite these advantages, concerns have arisen regarding artifacts introduced tagging systems presence lysine residues on them, prompting development alternative tools. In study, we introduce HiBiT-RR nLuc

Language: Английский

Lysineless HiBiT and NanoLuc Tagging Systems as Alternative Tools for Monitoring Targeted Protein Degradation DOI Creative Commons
Hanfeng Lin, Kristin M. Riching,

May Poh Lai

et al.

ACS Medicinal Chemistry Letters, Journal Year: 2024, Volume and Issue: 15(8), P. 1367 - 1375

Published: July 28, 2024

Target protein degradation (TPD) has emerged as a revolutionary approach in drug discovery, leveraging the cell's intrinsic machinery to selectively degrade disease-associated proteins. Nanoluciferase (nLuc) fusion proteins and NanoBiT technology offer two robust sensitive screening platforms monitor subtle changes abundance induced by TPD molecules. Despite these advantages, concerns have arisen regarding potential artifacts introduced tagging systems due presence of lysine residues on them, prompting development alternative tools. In this study, we introduce HiBiT-RR nLucK0, variants devoid residues, mitigate such artifacts. Our findings demonstrate that maintains similar sensitivity binding affinity with original HiBiT. Moreover, comparison between nLucWT nLucK0 constructs reveals variations patterns certain molecules, emphasizing importance choosing appropriate ensure reliability experimental outcomes studying processes.

Language: Английский

Citations

10
Computational approaches to aid PROTAC drug discovery DOI

Sohini Chakraborti,

Kirsten McAulay

Elsevier eBooks, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 1, 2025

Language: Английский

Citations

0
Interplay of PROTAC Complex Dynamics for Undruggable Targets: Insights into Ternary Complex Behavior and Linker Design DOI
Harish Kumar,

M. Elizabeth Sobhia

ACS Medicinal Chemistry Letters, Journal Year: 2024, Volume and Issue: 15(8), P. 1306 - 1318

Published: July 29, 2024

Protein degraders, such as bifunctional proteolysis-targeting chimeras (PROTACs), selectively eliminate target proteins by leveraging the natural protein degradation machinery. PROTACs bridge with an E3 ligase, which induces ubiquitination and degradation. Investigating ternary complex structures elucidates molecular mechanisms of their formation This study examines binding dynamics ligases (VHL, CRBN, cIAP) interest, focusing on dynamics, cooperativity, selectivity, linker length, PROTAC conformations. The influence interface residues lengths specific conformations for is highlighted. Utilizing steered simulations, provides comprehensive parameters behavior stability diverse complexes. These insights are crucial designing targeting disease-causing advancing development degradable complexes therapeutic applications.

Language: Английский

Citations

3
Extrapolating Lessons from Targeted Protein Degradation to Other Proximity-Inducing Drugs DOI Creative Commons
Georg E. Winter

ACS Chemical Biology, Journal Year: 2024, Volume and Issue: unknown

Published: Sept. 12, 2024

Targeted protein degradation (TPD) is an emerging pharmacologic strategy. It relies on small-molecule "degraders" that induce proximity of a component E3 ubiquitin ligase complex and target to ubiquitination subsequent proteasomal degradation. Essentially, degraders thus expand the function ligases, allowing them degrade proteins they would not recognize in absence small molecule. Over past decade, insights gained from identifying, designing, characterizing various have significantly enhanced our understanding TPD mechanisms, precipitating rational degrader discovery strategies. In this Account, I aim explore how these can be extrapolated anticipate both opportunities challenges utilizing overarching concept proximity-inducing pharmacology manipulate other cellular circuits for dissection biological mechanisms therapeutic purposes.

Language: Английский

Citations

3
Mechanistic insights into a heterobifunctional degrader-induced PTPN2/N1 complex DOI Creative Commons
Qi Hao, Manoj K Rathinaswamy,

Kelly L. Klinge

et al.

Communications Chemistry, Journal Year: 2024, Volume and Issue: 7(1)

Published: Aug. 16, 2024

PTPN2 (protein tyrosine phosphatase non-receptor type 2, or TC-PTP) and PTPN1 are attractive immuno-oncology targets, with the deletion of Ptpn1 Ptpn2 improving response to immunotherapy in disease models. Targeted protein degradation has emerged as a promising approach drug challenging targets including phosphatases. We developed potent PTPN2/N1 dual heterobifunctional degraders (Cmpd-1 Cmpd-2) which facilitate efficient complex assembly E3 ubiquitin ligase CRL4CRBN, mediate cells mice. To provide mechanistic insights into cooperative formation introduced by degraders, we employed combination structural approaches. Our crystal structure reveals how is recognized tri-substituted thiophene moiety degrader. further determined high-resolution DDB1-CRBN/Cmpd-1/PTPN2 using single-particle cryo-electron microscopy (cryo-EM). This that degrader induces proximity between CRBN PTPN2, albeit large conformational heterogeneity this ternary complex. The molecular dynamic (MD)-simulations constructed based on cryo-EM exhibited rigid body movement illustrated interactions CRBN. Together, our study demonstrates development potential applications cancer immunotherapy. Furthermore, workflow could help understand nature degrader-induced complexes. 2) however, targeting poses significant challenges. Here, authors report reveal biochemical structures cereblon X-ray diffraction, MD simulations.

Language: Английский

Citations

1
Protein Degradation in Focus DOI
David Zollman, Kirsten McAulay

Nature Chemical Biology, Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 29, 2024

Citations

1
Lysineless HiBiT and NanoLuc Tagging Systems as Alternative Tools Monitoring Targeted Protein Degradation DOI Open Access
Hanfeng Lin, Kristin M. Riching,

May Poh Lai

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: May 17, 2024

Target protein degradation (TPD) has emerged as a revolutionary approach in drug discovery, leveraging the cell's intrinsic machinery to selectively degrade disease-associated proteins. Proteolysis-Targeting Chimeras (PROTACs) exemplify this strategy, exploiting heterobifunctional molecules induce ubiquitination and subsequent of target The clinical advancement PROTACs underscores their potential therapeutic intervention, with numerous projects progressing through stages. However, monitoring subtle changes abundance induced by TPD demands highly sensitive assays. Nano-luciferase (nLuc) fusion proteins, or NanoBiT technology derived from it, offer robust screening platform due high sensitivity stability. Despite these advantages, concerns have arisen regarding artifacts introduced tagging systems presence lysine residues on them, prompting development alternative tools. In study, we introduce HiBiT-RR nLuc

Language: Английский

Citations

0