Structural Basis of Inhibition of DCLK1 by Ruxolitinib
Abstract
:1. Introduction
2. Results
2.1. Screening with Clinical Library Reveals Small-Molecule Doublecortin-Like Kinase 1 (DCLK1) Binders
2.2. Ruxolitinib Shows Significant off Target Activity against DCLK1
2.3. Overall Structural Features of the Ruxolitinib-Bound DCLK1
2.4. Dimerization of the Non-Phosphorylated DCLK1
2.5. Interaction Mode of Ruxolitinib with DCLK1
2.6. Comparison of the Binding Mode of Ruxolitinib with Other DCLK1 Inhibitors and ATP
3. Discussion
4. Materials and Methods
4.1. Cloning, Protein Expression, and Purification
4.2. Differential Scanning Fluorimetry (DSF) Assay
4.3. HTRF Assay
4.4. Surface Plasmone Resonance (SPR) Analysis of DCLK1
4.5. Crystallization, Data Collection, and Structure Determination
4.6. Molecular Docking Simulation
4.7. Molecular Dynamics Simulation
4.8. Data Deposition
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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PDB Entry | 7F3G |
---|---|
Data collection | |
Diffraction source | SPring-8 BL44XU |
Wavelength (Å) | 1.0000 |
Temperature (K) | 100 |
Space group | P21 |
a, b, c (Å) | 57.248, 80.118, 59.338 |
α, β, γ (°) | 90.000, 90.662, 90.000 |
Resolution range (Å) a | 47.68–2.09 (2.21–2.09) |
Total no. of reflections a | 118,719 (17,485) |
No. of unique reflections a | 59,261 (9116) |
Completeness (%) a | 95.5 (90.8) |
Redundancy a | 2.0 (1.9) |
〈I/σ(I)〉 a | 8.41 (1.04) |
Rsyma | 0.050 (0.725) |
Rmeasa | 0.067 (0.982) |
CC1/2a | 0.998 (0.622) |
Model refinement | |
Overall B factor from Wilson plot (Å2) | 44.0 |
No. of reflections, working set a | 29,588 (2100) |
No. of reflections, test set a | 1568 (131) |
final Rwork b | 0.210 |
final Rfree b | 0.253 |
No. of non-H atoms | 4333 |
protein | 4236 |
ruxolitinib | 46 |
water | 51 |
Average B factors (Å2) | 67.04 |
protein | 67.27 |
ruxolitinib | 61.19 |
water | 52.89 |
Root-mean-square deviations | |
bonds (Å) | 0.0048 |
angles (°) | 1.1384 |
Ramachandran plot c | |
favored (%) | 97.94 |
allowed (%) | 2.06 |
Rotamer c | |
favored (%) | 98.29 |
allowed (%) | 1.71 |
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Jang, D.M.; Lim, H.J.; Hahn, H.; Lee, Y.; Kim, H.K.; Kim, H.S. Structural Basis of Inhibition of DCLK1 by Ruxolitinib. Int. J. Mol. Sci. 2021, 22, 8488. https://doi.org/10.3390/ijms22168488
Jang DM, Lim HJ, Hahn H, Lee Y, Kim HK, Kim HS. Structural Basis of Inhibition of DCLK1 by Ruxolitinib. International Journal of Molecular Sciences. 2021; 22(16):8488. https://doi.org/10.3390/ijms22168488
Chicago/Turabian StyleJang, Dong Man, Hyo Jin Lim, Hyunggu Hahn, Yeon Lee, Hark Kyun Kim, and Hyoun Sook Kim. 2021. "Structural Basis of Inhibition of DCLK1 by Ruxolitinib" International Journal of Molecular Sciences 22, no. 16: 8488. https://doi.org/10.3390/ijms22168488