FLUORESCENCE DETECTED CIRCULAR DICHROISM OF DINUCLEOSIDE PHOSPHATES. A STUDY OF SOLUTION CONFORMATIONS AND THE TWO-STATE MODEL

Three fluorescent dinucleoside phosphates containing 1,N6-ethenoadenosine (eA), eApeA, eApeC, and eApU were studied using fluorescence-detected circular dichroism (FDCD), circular dichroism (CD), and absorption measurements. The FDCD data indicate that eApeC and eApU can be described as two-state systems consisting of a fluorescent species and a stacked, nonfluorescent species. Thermodynamic stacking parameters are calculated for these molecules using the van't Hoff equation. eApeA is found to be a more complicated system with a fluorescent CD which is different in shape, but comparable in magnitude, to the conventional CD of the dimer. This molecule, unlike the other two dinucleoside phosphates, cannot be characterized as a two-state system; it is described as consisting of at least three states at temperatures above 35°C. The CD data were subjected to a linear analysis in order to determine the minimum number of states present. In agreement with the FDCD data, eApeC and eApU are found to consist of a minimum of two states, while eApeA is indicated to have at least three. The more complicated behavior of the latter dimer is also indicated by the values of the unstacked CD obtained in the van't Hoff analysis.