The binding of Mn2+, Ca2+, and rare earth ions to apoconcanavalin A has been studied by water proton relaxation enhancement, electron paramagnetic resonance spectroscopy, and fluorescence spectroscopy. An electron paramagnetic resonance and water proton relaxation rate study of the titration of apoconcanavalin A with Mn2+ gives evidence of two equivalent binding sites per monomer with KD = 50 μm ± 4 μm. When a similar Mn2+ titration of apoconcanavalin A is performed in the presence of Ca2+ ion, very little free Mn2+ is detected by electron paramagnetic resonance until the two Mn2+ binding sites per monomer are filled. The substitution of a rare earth ion for Ca2+ ion in the above experiment often resulted in a slight displacement of Mn2+ from the transition metal site as detected by electron paramagnetic resonance. A water proton relaxation rate study of the titration of apoconcanavalin A with Gd3+ reflects two binding sites with a KD = 40 μm ± 4 μm and two with a KD = 200 μm ± 50 μm. The fluorescence emission spectrum of concanavalin A (λem = 340 nm) is slightly quenched by the addition of Tb3+ while Tb3+ fluorescence is greatly enhanced. A fluorometric titration of apoconcanavalin A with Tb3+ also reflects two sites with a KD = 40 μm ± 15 μm and two with a KD = 270 μm ± 50 μm.
ASJC Scopus subject areas
- Molecular Biology