| Lectin is a kind of protein or glycoprotein, which binds specifically andirreversibly monosaccharides or oligosaccharides. It is widely distributed in organismof all kinds in nature and plays important physiological roles. For example, as arecognizing molecule, it functions in the infection of viruses, bacteria and parasites;in the targeting of cells and soluble constituents; and in the metastasis, growth anddifferentiation of cancer cells. In this paper, the methods of SDS-PAGE, fluorescenceand circular dichroism are adopted to investigate the stability of Dolichos purpureuslectin (DPL) and the relationship between its structure and functions; thus the workhere lays a good foundation for further study of the structure of higher order andphysiological roles of DPL.In this paper, SDS-PAGE was adopted to investigate the stability of DPL. Theresult showed that in the temperature range of 35℃—65℃, DPL molecules did notaggregate, a small part of DPL molecules aggregated after being incubated at 75℃,over half of the DPL molecules aggregated after being incubated at 85℃, and almostall the DPL molecules aggregated after being incubated at 95℃.The stability of DPL was also investigated by using the hydrophobic probe 1,8-anilinonaphthalene sulfonic acid (ANS). It was found that in the temperature 35℃—75℃, the fluorescence intensity of ANS was very low, indicating the conformationof DPL did not change and no hydrophobic site was revealed, the fluorescenceintensity of ANS was substantially increased after being incubated at 85℃,suggesting that some hydrophobic sites were revealed, and DPL underwent thoroughinactivation after being incubated at 95℃. The measurement of intrinsic fluorescence was also carried out to study thevariation of DPL conformation after being incubated with different concentrations ofmannose and fructose. The results showed that the position of emission peak did notchange after being incubated with different concentrations of mannose, indicating themicroenvironment of tryptophan residues did not change. While mannose below theinhibition concentration led to the ordering of DPL conformation and thereforelowered the fluorescence intensity of DPL. Mannose over the inhibition concentrationfacilitated the transformation of the equilibrium of DPL-mannose binding to thebinding of mannose and consequently favored the lowering of DPL fluorescenceintensity.The fluorescence measurement of DPL incubated with fructose showed thatfructose did not influence the conformation of DPL, and the decrease of DPLfluorescence intensity was due to the static quenching caused by fructose.The secondary structures of DPL were investigated by measuring circulardichroism (CD) of DPL treated under different conditions. The maintenance of CDshape and signal intensity after being treated with urea under the concentration of 4mol/L suggested that DPL under these conditions sustained its secondary structures.After being treated with 8 mol/L urea, DPL was reduced in signal intensity withsimilar CD shape, indicating DPL maintained a certain degree of secondary structures.The CD results of DPL at different pH values showed that too alkaline or too acidicconditions both essentially influenced the content of secondary structures. Mannoseover the inhibition concentration did not necessarily affect the secondary structures ofDPL, and mannose below the inhibition concentration caused the increase of contentfα-helices andβ-strands, while fructose did not affect the secondary structures ofDPL. The CD spectra of DPL incubated at different temperatures showed that thecontent of secondary structures began to change after being incubated at 75℃, and itunderwent an obvious change after being incubated at an temperature over 85℃. Thecalculation results of secondary structure contents indicated that DPL contain a highcfntent fβ-strands and a low cfntent fα-helices, the latter decreased with theincrease of temperature at which DPL was incubated. The profile of the change of α-helices suggested that the unfolding of DPL was a two-state process.
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