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Updated: Jan 30, 2026
Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays
Published on: October 23, 2019
Elucidating the Role of Protein-Protein Interactions in Modulating Inhibitor Affinity and Release Mechanisms in
Shreya Mukherjee1, Niladri Patra1
1Department of Chemistry and Chemical Biology, Indian Institute of Technology (ISM) Dhanbad, Dhanbad 826004, India.
Abstract:
Protein-protein (pp) interactions make up a varied class of potential therapeutic targets for malignancy, which cause disturbances in the alternating splicing of SR proteins by SRPK due to nuclear accumulation. SRPK1 phosphorylates several serine residues in the RS domain of ASF/SF2, a classic SR protein. Substrate selectivity is dependent on protein interactions beyond the kinase active site. The RS domain's phosphorylation cycle needs strong, sustained SRPK-SR binding. In light of this evidence, the present research investigates the dynamics of protein-protein complexes bound to nucleotide triphosphates and inhibitor molecules. The influence of protein-protein interactions and the role of small molecules at the binding site, exerting competitive inhibition, were examined using classical molecular dynamics simulations complemented by statistical analysis. Trajectory visualization and analysis revealed differential ASF-SRPK binding in the presence of ATP and MSC1186 (the most recent SRPK inhibitor). In fact, conformational changes and binding orientations of small molecules, together with ASF-SRPK interactions, are interdependent in sustaining biological functions. Free energy of binding of the small molecules at the active pocket was empirically obtained from the Generalized Born Implicit Solvation Model with details of the residue-wise contribution toward binding. A precise absolute binding free energy approach, based on streamlined alchemical free energy perturbation, was applied to evaluate the binding affinity at the competitive pocket and yielded results consistent with experimental data. Enhanced sampling method─"random accelerated molecular dynamics (RAMD)", including PMF evaluation and path analysis─was employed to explore the egression route and investigate the exit dynamics of small molecules from the binding pocket. This study lays the groundwork for novel therapeutic design methodologies while improving our molecular-level understanding of protein-protein-small-molecule interactions linked to carcinogenesis.
Insights
This study reveals how small molecules like MSC1186 interact with ASF-SRPK protein complexes, offering new insights into cancer therapeutics by understanding these crucial protein-protein interactions.
Area of Science:
- Biochemistry and Molecular Biology
- Computational Chemistry
- Pharmacology
Background:
- Protein-protein interactions are key therapeutic targets in cancer, particularly involving SR protein kinase 1 (SRPK1) and ASF/SF2.
- Dysregulated SRPK1 activity and aberrant splicing of SR proteins contribute to carcinogenesis.
- Understanding the dynamics of SRPK-SR protein binding and the role of inhibitors is crucial for therapeutic development.
Purpose of the Study:
- To investigate the molecular dynamics of protein-protein complexes involving SRPK1 and ASF/SF2 in the presence of nucleotide triphosphates and inhibitor molecules.
- To elucidate the binding mechanisms and affinities of small molecule inhibitors, such as MSC1186, at the ASF-SRPK active site.
- To explore the egress dynamics of small molecules from the binding pocket using enhanced sampling methods.
Main Methods:
- Classical molecular dynamics simulations and statistical analysis.
- Generalized Born Implicit Solvation Model for binding free energy calculations.
- Alchemical free energy perturbation and Random Accelerated Molecular Dynamics (RAMD) for binding affinity and egress pathway analysis.
Main Results:
- Differential binding of ASF-SRPK was observed with ATP and the inhibitor MSC1186.
- Conformational changes and small molecule binding orientations are interdependent with ASF-SRPK interactions.
- Binding free energy calculations using advanced computational methods showed consistency with experimental data.
- RAMD simulations provided insights into the exit dynamics of small molecules from the binding pocket.
Conclusions:
- The study provides a detailed molecular-level understanding of protein-protein-small-molecule interactions in the context of carcinogenesis.
- Findings support the development of novel therapeutic strategies targeting SRPK1-mediated aberrant splicing.
- The computational methodologies employed offer a robust framework for future drug design and optimization.
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