The immune response to soluble D10 TCR: analysis of antibody and T cell responses

Interactions between short single-stranded DNA ligands and fluorescent DNA indicator dyes were used to investigate binding selectivity of the ligands. Conformational differences among four DNA ligands of different sequence and structure, including two that form a G-quartet and two that do not, were confirmed by circular dichroism spectroscopy. Their interactions with indicator dyes YO-pro-1 iodide (YO) and YOYO-1 iodide (YOYO) were probed using measurements of dye absorbance; induced circular dichroism; and fluorescence spectra, anisotropy, and lifetime. Equilibrium binding constants and stoichiometry were determined as well. Results indicate significant differences among the dye interactions and binding stoichiometries of the four ligands. One of the G-quartet forming ligands, a 20-mer of sequence 5'-GGTTTTGGTTTTGGTTTTGG-3', shows distinctly different interactions from the other three ligands, all of which are 15-mers. These studies illustrate the importance of sequence and conformation in determining the binding interactions of short single-stranded DNA.