The Binding of Terbium Ions to Tubulin Induces Ring Formation

Abstract The intrinsic fluorescence excitation and emission spectra of chicken brain tubulin showed the characteristic tryptophan fluorescence. The emission spectrum of Tb 3+ in the presence of tubulin and GTP excited at 295 nm, showed four peaks, with the maxima at 490, 545, and 586 am and a minor peak around 620 nm. Titration of tubulin with Tb 3+ was followed by the increment in luminescence at 545 nm and showed a sigmoidal curve where the initial lag interval and the maximal luminescence intensity depended on tubulin concentration. The presence of Mg 2+ , Co 2+ , and Zn 2+ diminished both the sigmoidicity of the curve and the maximal luminescence intensity. Titration of tubulin with Tb 3+ also produced a sigmoidal increase in turbidity, which was shifted to the left with respect to the luminescence curve. The dependence of turbidity on the wavelength of the Tb 3+ -induced polymers revealed that the large structures formed were not microtubules. Electron microscopy of the aggregates induced by Tb 3+ showed mainly a lattice of double rings with side-by-side contacts. These results indicate that Tb 3+ induces principally double ring formation and that these rings (33 ± 2 am external diameter) aggregate in large-ordered arrays. The luminescence of Tb 3+ seems to be induced mainly by the aggregation of rings.