Formation and stability of nucleotide complexes:: Raman titration investigation

In order to monitor the formation of complexes between modified and wild type nucleotides, a concept based on spectroscopic titration combined with factor analysis data treatment has been developed. A set of samples containing two homopolynucleotides with complementary bases at various concentration ratios has been prepared. Other conditions, i.e. total nucleotide concentration, temperature, ionic strength, divalent ions percentage and pH, were conserved. The series of Raman and UV-absorption spectra obtained from these samples have been treated by factor analysis and the results then fitted to the equilibrium equations of 1 : 1 and 1 : 2 complexes. By this way, spectra of individual unpaired nucleotides, 1 : 1 and 1 : 2 complexes, and values of the stability constants can be determined at the same time. While the former characterise the structure of the individual species, the latter parameters implicate their percentage in each sample. The approach has been tested on a Poly(rA)-poly(rU) model system. (C) 1999 Elsevier Science B.V. All rights reserved.