Sequence and Environmental Effect on the Formation of G‐Triplex DNA

It has recently been shown that certain G‐quadruplex DNA (G4) structures fold via a “G‐triplex” intermediate, and that the corresponding truncated G‐quadruplex‐forming oligonucleotides also adopt G‐triplex structures. However, it is not clear how general G‐triplex formation is, or how sequence and buffer conditions affect G‐triplex formation. Our approach to investigate G‐triplex formation by a series of truncated G4‐DNA sequences used CD spectroscopy, UV thermal‐difference spectroscopy, CD derived and UV derived melting temperature measurements, and native PAGE gels to characterize the topology and thermal stability of these structures in Li + , Na + , K + , Mg 2+ and Ca 2+ containing buffers. G‐triplex formation is a general phenomenon for a wide range of truncated G4 DNA sequences. We examined 16 different variants of the G ≥2 T 1–4 G ≥2 T 1–4 G ≥2 sequence as well as truncated versions of the TBA and human telomeric G4 DNA in both forward and reverse permutations. All of these sequences can adopt G‐triplex structures. However, we note that the number of nucleotides in the loop and sequence direction affect G‐triplex topology. Sequences with longer G‐tracks tends to form parallel G‐triplex, as do sequences with fewer loop residues. Permutation of the direction of the truncated G4 DNA sequence also affect G‐triplex folding topology. Environmental effects on topology were also noted, with divalent metal ions (Mg 2+ and Ca 2+ ) favoring parallel topologies. Using thermal‐difference spectroscopy, we have identified two families of spectra that correlate to G‐triplexes with a parallel topology. Analysis by native PAGE indicates that some sequences may form multiple G‐triplex topologies under certain environmental conditions. Temperature melts have provided information regarding sequence composition, sequence direction, and environmental conditions on the thermal stability of G‐triplex DNA. Sequences with longer G‐runs form G‐triplex structures that are more thermally stable (GGGG > GGG > GG). Shorter T‐loops form G‐triplex structures that are more thermally stable (TT >TTT > TTTT). Direction of sequence also effects the stability. Environmental conditions such as the presence of different cations have an effect of stability (Ca 2+ > K + > Mg 2+ ≥ Na + ). The biological relevance of G‐triplex DNA is still not entirely understood. Understanding how sequence effects and environmental conditions affect G‐triplex structure and stability is essential for better studying G‐quadruplex DNA and the biological relevance of G‐triplex DNA. Support or Funding Information CPRIT HIHR Grant RP160852 and Texas State University. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .