Formation and stability of nucleotide complexes: Raman titration investigation

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