Studies of the binding mechanism between aptamers and thrombin by circular dichroism, surface plasmon resonance and isothermal titration calorimetry.

Thrombin, a multifunctional serine protease, has both procoagulant and anticoagulant functions in human blood. Thrombin has two electropositive exosites. One is the fibrinogen-binding site and the other is the heparin-binding site. Over the past decade, two thrombin-binding aptamers (15-mer and 29-mer) were reported by SELEX technique. Recently, many studies examined the interactions between the 15-mer aptamer and thrombin extensively, but the data on the difference of these two aptamers binding to thrombin are still lacking and worth investigating for fundamental understanding. In the present study, we combined conformational data from circular dichroism (CD), kinetics and thermodynamics information from surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) to compare the binding mechanism between the two aptamers with thrombin. Special attentions were paid to the formation of G-quadruplex and the effects of ions on the aptamer conformation on the binding and the kinetics discrimination between specific and nonspecific interactions of the binding. The results indicated reasonably that the 15-mer aptamer bound to fibrinogen-binding site of thrombin using a G-quadruplex structure and was dominated by electrostatic interactions, while the 29-mer aptamer bound to heparin-binding site thrombin using a duplex structure and was driven mainly by hydrophobic effects.

Lin PH ，Chen RH ，Lee CH ，Chang Y ，Chen CS ，Chen WY ... -  《-》

Surface plasmon resonance spectroscopy study of interfacial binding of thrombin to antithrombin DNA aptamers.

Tang Q ，Su X ，Loh KP 《JOURNAL OF COLLOID AND INTERFACE SCIENCE》

Coexistence of G-quadruplex and duplex domains within the secondary structure of 31-mer DNA thrombin-binding aptamer.

Dolinnaya NG ，Yuminova AV ，Spiridonova VA ，Arutyunyan AM ，Kopylov AM ... -  《-》

The circular dichroism and differential scanning calorimetry study of the properties of DNA aptamer dimers.

We have applied circular dichroism (CD), temperature-gradient gel electrophoresis (TGGE) and differential scanning calorimetry (DSC) to study the properties of novel bioengineered DNA aptamer dimers sensitive to fibrinogen (F) and heparin (H) binding sites of thrombin and compared them with canonical single stranded aptamer sensitive to fibrinogen binding site of thrombin (Fibri). The homodimer (FF) and heterodimer (FH) aptamers were constructed based on hybridization of their supported parts. CD results showed that both FF and FH dimers form stable guanine quadruplexes in the presence of potassium ions like those in Fibri. The thermal stability of aptamer dimers was slightly lower compared to those of canonical aptamers, but sufficient for practical applications. Both FF and FH aptamer dimers exhibited a potassium-dependent inhibitory effect on thrombin-mediated fibrin gel formation, which was on average two-fold higher than those of canonical single stranded Fibri aptamers.

Poniková S ，Tlučková K ，Antalík M ，Víglaský V ，Hianik T ... -  《-》

A circular dichroism study of the stability of guanine quadruplexes of thrombin DNA aptamers at presence of K+ and Na+ ions.

Poniková S ，Antalík M ，Hianik T 《-》