In vitro study of the binding of terbium to tryptophan by 1H-NMR and fluorescence spectroscopy.

Terbium (Tb), a rare earth element having an ionic radius similar to that of Ca2+, is used as an antagonist of Ca2+. Recently, we found that when giant axon membrane of a loligo treated with Tb was excited at 290 nm, where tryptophan (Trp) gives absorption mechanism, emits fluorescence of Tb at 490 nm and 545 nm in addition to that of Trp at 345 nm. The present study was undertaken to locate on the Trp molecule the interaction between Trp and Tb by carrying out in vitro binding experiments with Tb and Trp and the molecular mechanism was analyzed by 1H-NMR spectroscopy and fluorescence spectroscopy. Fluorescence spectroscopy showed that Tb and Trp are combined at a molecular ratio close to 1:1. 1H-NMR spectroscopy revealed that Tb interacts with the amino group at the alpha-position and the imido group of the indole ring of Trp. Since all amino groups of Trp are involved in peptide linkages in vivo, it seems likely that the binding of Tb to Trp is undertaken by the imido group of the indole ring.