Enhancement of Inhibitory Activity by Combining Allosteric Inhibitors Putatively Binding to Different Allosteric Sites on Cathepsin K
Abstract
:1. Introduction
2. Results
2.1. Identification of Cathepsin K Inhibitors in the Extract of Chamaecrista Nomame and Analysis of Specificity of Cathepsin K Inhibitors, including Pheophytin a Derivatives
2.1.1. Purification of Cat K Inhibitors from the Methanol Extract of Chamaecrista nomame
2.1.2. Identification of Two Types of Cat K Inhibitor in the Methanol Extract of Chamaecrista nomame and Their Inhibitory Activity towards Cat K and B
2.1.3. Preparation of Pheophytin a Derivatives and Their Inhibitory Activity towards Cat K and Cat B
2.2. Mode of Cathepsin K Inhibition by Pheophytin a and Pheophorbide b and Determination of Kinetic Parameters
2.2.1. Allosteric Inhibition of Cat K by Pheophytin a and Pheophorbide b
2.3. Analysis of Allosteric Binding of Pheophytin a and Pheophorbide b Using AutoDock Vina
2.3.1. Docking Simulation of Pheophytin a and Pheophorbide b to Cat K
2.3.2. Pheophytin a and Pheophorbide b Bind to Each Allosteric Site That Is Different from the Allosteric Sites on Cat K, to Which the Known Allosteric Inhibitors NSC13345 and NSC94914 Bind
2.4. Additional Inhibition of Cat K by a Combination of Allosteric Inhibitors That Bind to Different Allosteric Sites
3. Discussion
4. Materials and Methods
4.1. Chemical Reagents and Instruments
4.2. Preparation of Extract of Chamaecrista nomame and Purification of Cat K Inhibitors
4.3. Preparation of Pheophytin a Derivatives from Extract of Spinach
4.4. Measurement of Inhibitory Activity of Inhibitors against Cat K and Cat B
4.5. Kinetic Analysis of Cat K Inhibition
4.6. Docking Study
4.7. Inhibition of Cathepsin K by Combination of Allosteric Inhibitors
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Sample Availability
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Sato, S.; Yamamoto, K.; Ito, M.; Nishino, K.; Otsuka, T.; Irie, K.; Nagao, M. Enhancement of Inhibitory Activity by Combining Allosteric Inhibitors Putatively Binding to Different Allosteric Sites on Cathepsin K. Molecules 2023, 28, 4197. https://doi.org/10.3390/molecules28104197
Sato S, Yamamoto K, Ito M, Nishino K, Otsuka T, Irie K, Nagao M. Enhancement of Inhibitory Activity by Combining Allosteric Inhibitors Putatively Binding to Different Allosteric Sites on Cathepsin K. Molecules. 2023; 28(10):4197. https://doi.org/10.3390/molecules28104197
Chicago/Turabian StyleSato, Shun, Kana Yamamoto, Moeno Ito, Katsutoshi Nishino, Takanao Otsuka, Kazuhiro Irie, and Masaya Nagao. 2023. "Enhancement of Inhibitory Activity by Combining Allosteric Inhibitors Putatively Binding to Different Allosteric Sites on Cathepsin K" Molecules 28, no. 10: 4197. https://doi.org/10.3390/molecules28104197