Molecular Glue/PROTAC® Optimization Kit for RBM39-DCAF15

Products from BPS Bioscience require a minimum order value above 400€

Applications: • Identify and optimize Molecular Glues/PROTACs targeting RBM39.• Design novel molecules targeting DCAF15.• Directly compare the activity of different Molecular Glues/PROTACs.

Background: DCAF15 is the substrate-binding component of the E3 protein ligase complex DDB1-DDA1-CUL4-Rbx1, involved in the ubiquitination and proteasomal degradation of target proteins. Binding of DCAF15 to a substrate protein engages the E3 ligase activity of the complex and results in the ubiquitination and ultimate degradation of the protein substrate. One of the most important targets of DCAF15 is RBM39 (RNA-binding motif protein 39). RBM39 is a key factor in tumor-targeted mRNA and protein expression. Recent studies have revealed that RBM39 is the unexpected target of aryl sulphonamides, which act as molecular glues between RBM39 and the DCAF15-associated E3 ubiquitin ligase complex leading to selective degradation of the target. E7820 is an example of aryl sulfonamides that recruit RBM39 to Rbx-CUL4-DDA1-DDB1-DCAF15 E3 ligase complex, leading to its ubiquitination and degradation by the proteasome. Development of novel MG and PROTACS involved in the recruitment of RBM39 provides a framework for future efforts to utilize DCAF15 to degrade other proteins of interest.

Contraindications: The final concentration of DMSO in the assay should not exceed 1%.

Avoid green and blue dyes that absorb light in the AlphaScreen signal emission range (λ=520-620 nm), such as Trypan Blue.

Avoid the use of potent singlet oxygen quenchers such as sodium azide (NaN₃) or metal ions (Fe²⁺, Fe³⁺, Cu²⁺, Zn²⁺ and Ni²⁺).

The presence of >1% RPMI 1640 culture medium leads to a signal reduction due to the presence of excess biotin and iron in this medium. Media like MEM, which lacks these components, does not affect AlphaScreen assays.

Description: The Molecular Glue/PROTAC® Optimization Kit for RBM39-DCAF15 Binding is designed for the testing and profiling of Molecular Glues (MG)/PROTACs targeting the RBM39 (RNA-binding protein 39) and DCAF15 (DDB1 and CUL4 associated factor 15) interaction. The Molecular Glue/PROTAC® Optimization Kit for RBM39-DCAF15 Binding comes in a convenient AlphaLISA® format, with enough optimized assay and detection buffer, purified recombinant RBM39 (amino acids 245-530) and Rbx1/CUL4B/DDB1/DCAF15/DDA1 complex for 384 reactions. This kit also contains the molecular glue E7820 and Arginine as controls.

Figure 1. Schematic representation of the Molecular Glue/PROTAC® Optimization Kit for RBM39-DCAF15 Binding assay kit principle.
The Molecular Glue/PROTAC® of interest is incubated with DCAF15 complex and RBM39, bringing them in close proximity. DCAF15 contains a His-tag, which is recognized by the donor beads. RBM39 contains a GST tag that binds to the GSH- AlphaLISA™ acceptor beads. Upon excitation of the donor bead, a singlet oxygen is generated by the bead. The singlet oxygen excites the acceptor bead, which emits light proportionally to the level of interaction between RBM39 and DCAF15

Storage Stability: This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.

Warnings: Avoid freeze/thaw cycles

Biosafety Level: Not applicable (BSL-1)

References: Nunes, J., et al. 2019 ACS Med Chem Lett; 10(7): 1081-1085

• SKU: BPS82251
• Manufacturer: BPS Bioscience
• Manufacturer SKU: 82251
• Green Labware: No
• Package Unit: 384 reactions
• Quantity Unit: PAK