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Updated: Oct 8, 2025
An Optimized Quantitative Pull-Down Analysis of RNA-Binding Proteins Using Short Biotinylated RNA
Published on: February 17, 2023
Microscale thermophoresis analysis of the molecular interaction between small nuclear ribonucleoprotein polypeptide G
Lloyd Mabonga1, Priscilla Masamba2, Albertus Kotze Basson1
1Department of Biochemistry and Microbiology, University of Zululand KwaDlangezwa 3886, South Africa.
Abstract:
Regulatory core-splicing proteins are now becoming highly promising therapeutic targets for the development of anti-cancer drugs. SNRPG and RBBP6 are two good examples of regulatory core-splicing proteins involved in tumorigenesis and tumor development whose multi-functional role is primarily mediated by protein-protein interactions. Over the years, skepticism abutting from the two onco-proteins has been mounting. Suggestive evidence using yeast 2-hybrid technique observed possible involvement between SNRPG and the RING finger domain of RBBP6. However, the putative interaction remains elusive and yet to be characterized. In this study, we developed the first MST-based assay to confirm the interaction between SNRPG and the RING finger domain of RBBP6. The results demonstrated a strong binding affinity between SNRPG and the RING finger domain of RBBP6 with a KD in the low nanomolar concentration range of 3.1596 nM. The results are congruent with previous findings suggesting possible involvement between the two proteins in cancer-cell networks, thereby providing a new mechanistic insight into the interaction between SNRPG and the RING finger domain of RBBP6. The interaction is therapeutically relevant and represents a great milestone in the anti-cancer drug discovery space. Identification of small molecule inhibitors to modulate the binding affinity between the two proteins would therefore be a major breakthrough in the development of new PPI-focused anti-cancer drugs.
Insights
Regulatory core-splicing proteins SNRPG and RBBP6 are implicated in cancer. This study confirms their interaction, revealing a potential new target for anti-cancer drug development.
Area of Science:
- Molecular Biology
- Biochemistry
- Oncology
Background:
- Regulatory core-splicing proteins are emerging as key therapeutic targets in oncology.
- SNRPG and RBBP6 are implicated in tumorigenesis, with their functions mediated by protein-protein interactions.
- Previous evidence suggested a link between SNRPG and the RBBP6 RING finger domain, but this interaction remained uncharacterized.
Purpose of the Study:
- To confirm and characterize the interaction between SNRPG and the RBBP6 RING finger domain.
- To provide mechanistic insight into the role of this interaction in cancer-cell networks.
- To explore the therapeutic potential of targeting this protein-protein interaction for anti-cancer drug discovery.
Main Methods:
- Development of a MicroScale Thermophoresis (MST)-based assay.
- Quantitative analysis of the binding affinity between SNRPG and the RBBP6 RING finger domain.
Main Results:
- The study successfully confirmed the interaction between SNRPG and the RBBP6 RING finger domain.
- A strong binding affinity was demonstrated, with a KD value in the low nanomolar range (3.1596 nM).
- These findings support the involvement of SNRPG and RBBP6 in cancer-related cellular networks.
Conclusions:
- The confirmed interaction between SNRPG and the RBBP6 RING finger domain offers a novel mechanistic insight into cancer development.
- This interaction represents a therapeutically relevant target for the development of new anti-cancer drugs.
- Identifying small molecule inhibitors to modulate this binding affinity could lead to a breakthrough in protein-protein interaction-focused cancer therapies.
