Polarization of ligand fluorescence was used to study the binding of 4-methylumbelliferyl β-d-galactopyranoside (MeUmb-Galp) to Abrus precatorious agglutinin. The binding of the fluorescent sugar to the lectin led to considerable polarization of the MeUmb-Galp fluorescence, which was also quenched by about 30% on binding to the lectin. The binding of the fluorescent sugar was carbohydrate-specific, as evidenced by inhibition of both fluorescence polarization and quenching when lectin was preincubated with lactose. The association constant as determined by fluorescence polarization is 1.42 × 104 M−1 at 25° C and is in excellent agreement with those determined by fluorescence quenching (Ka= 1.51 × 104 M−1) and equilibrium dialysis (Ka= 1.62 × 104 M−1) at 25° C. The numbers of binding sites as determined by fluorescence polarization, quenching and equilibrium dialysis agree very well with one another, n being equal to 2.0 ± 0.05. The consistency between the association constant value determined by fluorescence polarization, quenching and equilibrium dialysis shows the validity of this approach to study lectin-sugar interaction.
No takes yet. Share an insight, caveat, or question.
Khan et al. (2005) studied this question.