A simple, label-free optical method for studies on the G-quadruplex/duplex competition inside duplex DNAs using a G-quadruplex-specific probe—TMPipEOPP

Abstract

G-rich sequences that might form G-quadruplexes are common in some regions of the human genome. Most of these G-rich sequences are embedded in flanking duplex-forming sequences and coexist with complementary C-rich sequences. Therefore, the competition between G-quadruplexes and the duplex structures in flanking duplex DNAs must be considered in the corresponding G-quadruplex studies. Based on the high specificity of a porphyrin derivative (TMPipEOPP) for G-quadruplexes over duplexes and single-stranded DNAs, a simple method was developed for studying the competition between G-quadruplexes and duplexes in flanking duplex structures. In the presence of a complementary sequence, the core G-rich region of a long DNA sequence tended to form a duplex under dilute conditions and form a G-quadruplex under molecular crowding conditions. The method could also be used to monitor conversion between G-quadruplexes and duplexes in real time. Under dilute conditions, G-rich sequences rapidly hybridized with complementary strands to form stable duplexes that did not disassociate with time. If the G-rich sequences have been folded into G-quadruplexes, addition of a complementary sequence promoted slow conversion from G-quadruplexes to duplexes. However, under molecular crowding conditions, stable G-quadruplexes formed regardless of the presence of complementary sequences and the formed G-quadruplexes did not disassociate with time. Changing solution conditions from dilute to molecular crowding promoted rapid structural conversion of G-rich sequences from duplexes to G-quadruplexes. This method could be an important tool for G-quadruplex studies.