Dissociation Kinetics of an Enzyme−Inhibitor System Using Single-Molecule Force Measurements

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• 作者：Essa Mayyas ; Margarida Bernardo ; Lindsay Runyan ; Anjum Sohail ; Venkatesh Subba-Rao ; Mircea Pantea ; Rafael Fridman ; Peter M. Hoffmann
• 刊名：Biomacromolecules
• 出版年：2010
• 出版时间：December 13, 2010
• 年：2010
• 卷：11
• 期：12
• 页码：3352-3358
• 全文大小：246K
• 年卷期：v.11,no.12(December 13, 2010)
• ISSN：1526-4602

文摘

We report on an improved method to interpret single molecule dissociation measurements using atomic force microscopy. We describe an easy to use methodology to reject nonspecific binding events, as well as estimating the number of multiple binding events. The method takes nonlinearities in the force profiles into account that result from the deformation of the used polymeric linkers. This new method is applied to a relevant enzyme−inhibitor system, latent matrix metalloprotease 9 (ProMMP-9, a gelatinase), and its inhibitor, tissue inhibitor of metalloproteases 1 (TIMP 1), which are important players in cancer metastasis. Our method provides a measured kinetic off-rate of 0.010 ± 0.003 s⁻¹ for the dissociation of ProMMP9 and TIMP1, which is consistent with values measured by ensemble methods.