Fluorescence quenching and time-resolved fluorescence studies on Momordica charantia (bitter gourd) seed lectin

Abstract

Chemical modification studies implicated tryptophan (Trp) residues in the sugar binding activity of Momordica charantia lectin (MCL). In the present study, the accessibility and environment of Trp residues in MCL were investigated by intrinsic fluorescence quenching and time-resolved fluorescence. The emission γ(max) of native MCL in the absence as well as in the presence of 0.1 M lactose was around 335 nm, which shifted to 365 nm in the presence of 8 M urea, suggesting that the Trp residues which are predominantly buried in the hydrophobic core of the native lectin get exposed to the aqueous environment upon denaturation. At a quencher concentration of 0.5 M, the extent of quenching observed for the native MCL with acrylamide, I- and Cs+ was 46%, 17% and 12%, respectively. In the presence of 0.1 M lactose this quenching was smaller, suggesting that the sugar ligand provides a partial protection to the Trp residues. In time-resolved fluorescence measurements, the decay curves could be fitted well to a biexponential function with the estimated life times 0.92 ns and 4.64 ns for the native protein and 1.15 ns and 5.1 ns in the presence of 0.1 M lactose. All these results are consistent with the involvement of Trp residues in the sugar-binding activity of MCL.