Photophysics and binding constant determination of the homodimeric dye BOBO-3 and DNA oligonucleotides.
The interactions between single- and double-stranded DNA and the trimethine cyanine homodimer dye, BOBO-3 (1,1'-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-thiazole)-2-methylidene]pyridinium tetraiodide), have been investigated in detail using absorption and steady-state and time-resolved fluorescence spectroscopy. The dye interacts with both single-stranded and double-stranded DNA, under a variety of conditions, with changes in its spectral characteristics. Our results indicated that the complex formed between BOBO-3 dye and DNA oligonucleotides could not be explained with a simple, single intercalation mechanism; therefore, different modes of interaction were proposed. By using time-resolved fluorescence methodology and in-depth analysis of the fluorescence decay traces, we obtained the contribution of the different forms of BOBO-3: free in solution, a low affinity, electrostatically driven interaction with DNA, and a full bis-intercalation mechanism within the DNA double helix. With this information, we applied the McGhee-Von Hippel theory for two overlapping, noncooperative binding modes to obtain equilibrium binding constants and the number of sites occupied for each binding mode. Binding constants for dye/dsDNA complexes in complete bis-intercalation and externally bound were (8.8 +/- 1.1) x 10(5) and (2.6 +/- 0.3) x 10(5) M(-1), respectively. The corresponding recovered number of base pairs covered were 5.9 +/- 0.2 and 3.5 +/- 0.5 sites for each mode.
Ruedas-Rama
Maria J
MJ
Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada, Spain.
Orte
Angel
A
Crovetto
Luis
L
Talavera
Eva M
EM
Alvarez-Pez
Jose M
JM
D002232:Carbocyanines / Q000737:chemistry*; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry*; D007364:Intercalating Agents / Q000737:chemistry*; D008958:Models, Molecular; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry*; D010777:Photochemistry; D013050:Spectrometry, Fluorescence
C098829:BOBO 3; D002232:Carbocyanines; D004277:DNA, Single-Stranded; D005456:Fluorescent Dyes; D007364:Intercalating Agents; D009841:Oligonucleotides; D004247:DNA