Characterization of Structure and Stability of Long Telomeric DNA G-Quadruplexes

Abstract

In the current study, we used a combination of gel electrophoresis, circular dichroism, and UV melting analysis to investigate the structure and stability of G-quadruplexes formed by long telomeric DNAs from Oxytricha and human, where the length of the repeat (n)=4 to 12. We found that the Oxytricha telomeric DNAs, which have the sequence (TTTTGGGG)n, folded into intramolecular and intermolecular G-quadruplexes depending on the ionic conditions, whereas human telomeric DNAs, which have the sequence (TTAGGG)n, formed only intramolecular G-quadruplexes in all the tested conditions. We further estimated the thermodynamic parameters of the intramolecular G-quadruplex. We found that thermodynamic stabilities of G-quadruplex structures of long telomeric DNAs (n=5 to 12) are mostly independent of sequence length, although telomeric DNAs are more stable when n=4 than when n>or=5. Most importantly, when n is a multiple of four, the change in enthalpy and entropy for G-quadruplex formation increased gradually, demonstrating that the individual G-quadruplex units are composed of four repeats and that the individual units do not interact. Therefore, we propose that the G-quadruplexes formed by long telomeric DNAs (n>or=8) are bead-on-a-string structures in which the G-quadruplex units are connected by one TTTT (Oxytricha) or TTA (human) linker. These results should be useful for understanding the structure and function of telomeres and for developing improved therapeutic agents targeting telomeric DNAs.