Evaluation of the structure–activity relationship of thrombin with thrombin bin...

• Author(s): Diculescu, Victor Constantin ; Chiorcea-Paquim, Ana-Maria ; Eritja, Ramon ; Oliveira-Brett, Ana Maria
• Date: 2011
• Persistent ID: http://hdl.handle.net/10316/17993
• Origin: Estudo Geral - Universidade de Coimbra
• Subject(s): Thrombin; Aptamer; Quadruplex; Guanine; AFM; Electrochemistry
• Document Type: Article
• Language: English

Description

The structure–activity relationship of the complex between thrombin and thrombin binding aptamers (TBA) was evaluated by differential pulse voltammetry at a glassy carbon electrode and atomic force microscopy at a highly oriented pyrolytic graphite electrode. The effects on the interaction with thrombin of TBA primary and secondary structures as well as of its folding properties in the presence of alkaline metals were investigated. The complex between thrombin and single stranded aptamers involved the coiling of the single stranded molecules around thrombin structure leading to the formation of a robust TBA–thrombin complex that maintains the symmetry and conformation of the thrombin molecule. Monitoring both thrombin and TBA oxidation peaks, showed that the thrombin oxidation peaks occur at more positive potentials after TBA–thrombin complex formation. In the presence of K+ ions, the aptamers fold into quadruplex structures that facilitate the interaction with thrombin molecules. The TBA–thrombin complex adsorbs at the surface with the aptamer quadruplex always in preferential contact with the surface, and the thrombin molecules on top of the aptamer quadruplex structure, thus being less accessible to the electrode surface leading to the occurrence of thrombin oxidation peaks at less positive potentials.