Identification of NEK7 inhibitors: structure based virtual screening, molecular docking, density functional theory calculations and molecular dynamics simulations

Article

作者: Alsubaie, A. SA. ; Aziz, Mubashir ; Mahmoud, K. H. ; Alrowaili, Z. A. ; Al-Buriahi, M. S. ; Siddique, Farhan ; Rehman, Hafiz Muzzammel ; Ejaz, Syeda Abida

NEK7 is a NIMA related-protein kinase that plays a crucial role in spindle assembly and cell division. Dysregulation of NEK7 protein leads to development and progression of different types of malignancies including colon and breast cancers. Therefore, NEK7 could be considered as an attractive target for anti-cancer drug discovery. However, few efforts have been made for the development of selective inhibitors of NIMA-related kinase but still no FDA approved drug is known to selectively inhibit the NEK7 protein. Dacomitinib and Neratinib are two Enamide derivatives that were approved for treatment against non-small cell lung cancer and breast cancer respectively. Drug repurposing is a time and cost-efficient method for re-evaluating the activities of previously authorized medications. Thus, the present research involves the repurposing of two FDA-approved medications via comprehensive in silico approach including Density functional theory (DFTs) studies which were conducted to determine the electronic properties of the Dacomitinib and Neratinib. Afterward, binding orientation of selected drugs inside NEK7 activation loop was evaluated through molecular docking approach. Selected drugs exhibited potential molecular interactions engaging important amino acid residues of active site. The docking score of Dacomitinib and Neratinib was -30.77 and -26.78 kJ/mol, respectively. The top ranked pose obtained from molecular docking was subjected to Molecular Dynamics (MD) Simulations for investigating the stability of protein-ligand complex. The RMSD pattern revealed the stability of protein-ligand complex throughout simulated trajectory. In conclusion, both drugs displayed inhibitory efficacy against NEK7 protein and provide a prospective therapy option for malignant malignancies linked with NEK7.

2021-12-01·Protein & Peptide Letters4区 · 生物学

Optimized Expression of Recombinant Human NIMA-Related Kinase 7 (NEK7) with A Higher Purity in Escherichia coli

4区 · 生物学

Article

作者: Zhang, Rui-Han ; Li, Xiao-Li ; Wang, Ting-Ting ; Ji, Xu ; Xiao, Wei-Lie ; Pu, Yu-Kun ; Zhang, Xing-Jie ; Zeng, Lin

Background:NIMA (never in mitosis, gene A) serine/threonine kinase 7 (NEK7) is a regulator of mitosis spindle in mammals and is considered as a drug target of inflammasome related inflammatory diseases. However, most commercially available or reported recombinant NEK7 proteins are either inactive or have low purity. These shortcomings limit the pharmacological studies and development of NEK7 inhibitors.Objective:To elucidate what causes the NEK7 low purity in E. coli, and optimize a protocol to improve the protein purity.Methods:A comparative study of expression full length NEK7 with an N-terminal His-tag or a Cterminal His-tag was performed. His-affinity resin, ion exchange and gel filtration chromatography were used to purify NEK7. The protein was identified by mass spectrometry. The activity and folding of NEK7 were evaluated by chemiluminescent assay and thermal shift assay.Results:Our results demonstrated that N-terminal tagged protein was toxic to E. coli, resulting in incomplete translated products. The C-terminal tagged NEK7-His6 had a much higher purity than that of an N-terminal tag. The Ni2+ resin one-step purification led to a purity of 91.7%, meeting the criteria of most kinase assays. With two-step and three-step procedures, the protein purities were 94.7% and ~100%, respectively. The NEK7 purified in this work maintained its kinase activity and correct conformation, and the compound-protein interaction ability.Conclusion:Our optimized protocol could produce good purity of His tagged NEK7 in E. coli, and the kinase activity and biophysical characteristics of which are preserved.

2020-04-30·Biochemical Journal3区 · 生物学

Nek7 conformational flexibility and inhibitor binding probed through protein engineering of the R-spine

3区 · 生物学

Article

作者: Nasir, Nazia ; Mas-Droux, Corine ; Bayliss, Richard ; Yeoh, Sharon ; Byrne, Matthew J. ; Basmadjian, Christine ; Cunnison, Rory F. ; Cano, Céline ; Bhatia, Chitra ; Carr, Katherine H.

Nek7 is a serine/threonine-protein kinase required for proper spindle formation and cytokinesis. Elevated Nek7 levels have been observed in several cancers, and inhibition of Nek7 might provide a route to the development of cancer therapeutics. To date, no selective and potent Nek7 inhibitors have been identified. Nek7 crystal structures exhibit an improperly formed regulatory-spine (R-spine), characteristic of an inactive kinase. We reasoned that the preference of Nek7 to crystallise in this inactive conformation might hinder attempts to capture Nek7 in complex with Type I inhibitors. Here, we have introduced aromatic residues into the R-spine of Nek7 with the aim to stabilise the active conformation of the kinase through R-spine stacking. The strong R-spine mutant Nek7SRS retained catalytic activity and was crystallised in complex with compound 51, an ATP-competitive inhibitor of Nek2 and Nek7. Subsequently, we obtained the same crystal form for wild-type Nek7WT in apo form and bound to compound 51. The R-spines of the three well-ordered Nek7WT molecules exhibit variable conformations while the R-spines of the Nek7SRS molecules all have the same, partially stacked configuration. Compound 51 bound to Nek2 and Nek7 in similar modes, but differences in the precise orientation of a substituent highlights features that could be exploited in designing inhibitors that are selective for particular Nek family members. Although the SRS mutations are not required to obtain a Nek7–inhibitor structure, we conclude that it is a useful strategy for restraining the conformation of a kinase in order to promote crystallogenesis.

100 项与 NEK7 Targeting Program (Monte Rosa) 相关的药物交易

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