Characterization of the interaction of galectin-1 with sodium arsenite.
We previously showed that galectin-1 (GAL1) is an arsenic-binding protein. In the current study, we further characterize the interaction of GAL1 with sodium arsenite (As(III)). The GALl-As(III) complex was prepared from the cell extracts of GAL1-transfected Escherichia coli (E. coli) that were pretreated with As(III). The results of the circular dichroism (CD) spectrum of GAL1-As(III) exhibited a negative signal at around 205-210 nm, whereas that of GAL1 showed a negative signal at around 215-220 nm. This shift in the CD spectrum is indicative of a substantial change in the secondary structure arising from the binding of As(III) to the GAL1 protein. The UV absorptive spectrum of the GAL1-As(III) complex was significantly lower than that of GAL1 itself. A mobility shift binding assay showed that the GAL1-As(III) complex migrated closer than GAL1 toward the anode. Capillary electrophoretic analysis also showed that As(III) binding decreased the mobility of GAL1. These results further confirmed the structural change of the GAL1 complex with As(III). Furthermore, isothermal titration microcalometric studies showed that As(III) titration into the GAL1 protein solution was an endothermic process with absorption enthalpy (DeltaH(abs)) around 8-10 kJ/mol As(III). The affinity constant (K(d)) of As(III) toward GAL1 was around 8.239 +/- 2.627 microM as estimated by tryptophan (Trp) fluorescence quenching. However, the binding of As(III) did not significantly affect the biological activity of GAL1, since the GAL1-As(III) complex only partially lost its lectin activity. In addition, we show that GAL1-transfected KB cells accumulated more arsenic than did the parental cells. Taken together, these results suggest that GAL1 might serve as a target protein of As(III) in vivo, and the binding of GAL1 with As(III) could interfere with the excretion of As(III).
Department of Life Science, College of Science, National Central University & University System of Taiwan, Chung-Li, Taoyuan, Taiwan 32054, ROC.
Chen-Huan
CH
Lin
Chi-Fei
CF
Huang
Wen-Yih
WY
Chen
Yu-Ying
YY
Chang
Wang-Hsien
WH
Ding
Ming-Shen
MS
Lin
Szu-Hsun
SH
Wu
Rong-Nan
RN
Huang
D018053:Arsenites / Q000737:chemistry*; D002151:Calorimetry; D002942:Circular Dichroism; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D037483:Galectin 1 / Q000737:chemistry* / Q000235:genetics; D006358:Hot Temperature; D006801:Humans; D007202:Indicators and Reagents; D007624:KB Cells; D037102:Lectins; D017670:Sodium Compounds / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D014162:Transfection
D018053:Arsenites; D037483:Galectin 1; D007202:Indicators and Reagents; D037102:Lectins; D017670:Sodium Compounds; C017947:sodium arsenite