Interaction of RNase A with VO3- and VO2+ ions. Metal ion binding mode andprotein secondary structure.

Some of vanadyl complexes have shown potential to inhibit RNase activity by acting as transition state analogue, while at the same time not inhibiting DNase. To gain an insight into the interaction of protein with vanadate (V O3-) and vanadyl (VO2+) ions, the present study was designed to examine the binding of ribonuclase A (RNase A) with NaVO3 and VOSO4 in aqueous solutio n at physiological pH with metal ion concentrations of 0.001 mM to 1 mM, an d protein concentration of 2% w/v. Absorption spectra and Fourier transform infrared (FTIR) spectroscopy with self-deconvolution and second derivative resolution enhancement were used to determine the cation binding mode, ass ociation constant and the protein secondary structure in the presence of va nadate and vanadyl ions in aqueous solution. Spectroscopic results show that an indirect metal ion interaction occurs wi th the polypeptide C = O, C-N (via H2O) with overall binding constants of K (VO3-) = 3.93 x 10(2) M-1 and K(VO2+) = 4.20 x 10(3) M-1. At high metal ion concentrations, major protein secondary structural changes occur from that of the alpha-helix 29% (free enzyme) to 23-24%; beta-sheet (pleated and an ti) 50% (free enzyme) to 64-66% and turn 21% (free enzyme) to 10-12% in the metal-RNase complexes. The observed structural changes indicate a partial protein unfolding in the presence of high metal ion concentration.