Aminoquinolines: Fluorescent sensors to DNA - A minor groove probe. Experimental and in silico studies

An aminoquinoline (AQ4) was developed and proven to be a new and efficient DNA minor groove fluorescent probe. The specificity for DNA minor groove was attested by comparing it with well-established DNA probes such as Hoechst stain, acridine orange, and ethidium bromide. AQ4 was similar to the Hoechst stain, a classic minor groove probe, and opposite to acridine orange and ethidium bromide, the typical intercalating probes. An advantage of AQ4 to the Hoechst stain was the higher fluorescent signal-to-noise ratio (+DNA/-DNA). The interaction with DNA leads to an exclusive fluorescent band centered at 590 nm. The red-shifted fluorescent band is associated with a new absorption band (490 nm), revealing a ground-state complex formation. The complexation was also evidenced by circular dichroism, anisotropy, and fluorescence lifetime. The complex AQ4DNA was pH dependent, being favored in an acidic medium. This feature was related to the preferential interaction with a protonated form of AQ4, i.e., AQ4(H+), revealing the role of electrostatic forces, which was corroborated by the strong dependence on the ionic strength of the medium and, particularly, on magnesium ions. The complexes were studied by docking and molecular dynamics and confirmed the stability of AQ4(H+). Additionally, 500 ns simulations were performed by adding salts. The presence of the salts leads to the loss of the binding of AQ4(H+) after 52 ns (NaCl) and 142 ns (MgCl2). Experimental and in silico outcomes showed the advantages of the aminoquinoline over the commercial DNA minor groove stain, the Hoechst dye. Hence, we propose its further application in cell-based assays.

Automated summary

An aminoquinoline probe AQ4 has been developed as a new and efficient DNA minor groove fluorescent probe, demonstrating specificity for the minor groove compared to other established probes. AQ4 showed similar properties to Hoechst stain, but different from acridine orange and ethidium bromide. It had a higher fluorescent signal-to-noise ratio compared to Hoechst stain.