METAL-ION BINDING-PROPERTIES OF THE TRANSFERRINS - A V-51 NMR-STUDY

Transferrins can bind a wide range of di- and trivalent metal ions. Th ey have a bilobal structure where each domain contains a deep cleft th at binds a metal ion along with a synergistic anion. In this work, the binding of vanadate as VO2+ to the transferrins was studied by V-51 q uadrupolar central transition (QCT) NMR. Information about differences in the symmetry and motion of the bound metal ion was obtained from c hemical shift and line width differences for serotransferrin (sTf), la ctoferrin (lTf), and ovotransferrin (oTf). The effects of pH, ionic st rength, and temperature on the V-51 QCT NMR spectra of the bound VO2cations showed that the N-lobe binding site of sTf is unique as compar ed to the other proteins. Properties of the quadrupolar central transi tion were also investigated, revealing that temperature, magnetic held strength, and NMR pulse angle all induce predictable changes on the s econd-order dynamic frequency shift, spectral line width, and optimal pulse angle in the V-51 NMR spectra. Analysis of NMR spectra of V(V)(2 )-oTf and V(V)(2)-sTf at three magnetic fields allowed an estimation o f the quadrupolar coupling constants for these binding sites. This ind icates that the degree of coordination symmetry in the binding sites i s as follows: sTf N < sTf C < oTf N, C. Carbon-13 NMR studies revealed that VO2+ binding, in contrast to di- and trivalent metal ions, has n o requirement for a synergistic anion.