稀土离子与乳铁蛋白结合的光谱研究

Spectroscopic Studies on the Binding of Lanthanides to the Apolactoferrin

用紫外示差光谱、荧光光谱及圆二色谱等方法研究了 Tb3+ 和 Eu3+ 在 p H 7.4的条件下与乳铁蛋白及脱铁乳铁蛋白的结合作用 .结果表明 ,Tb3+及 Eu3+可特异性地结合在脱铁乳铁蛋白的两个 Fe3+结合部位 ,但不能从已经结合铁的乳铁蛋白中把铁置换出来 .测得 Tb3+与这两个部位结合的条件平衡常数为 lg K1 =8.4 8± 0 .2 4和 lg K2 =6.72± 0 .1 8( 2 5℃ ,0 .1 0 mol/L Na Cl,0 .1 0 mol/L Hepes,p H=7.4 ) . Tb3+ 在这两个位点结合时 ,蛋白质的构象发生变化 .在 Tb3+ 与蛋白质的浓度比低时 ,构象趋于紧缩 ,色氨酸残基进入疏水的环境 ;当 Tb3+ 结合得较多时 ,构象转而开放 ,色氨酸残基转向亲水性环境 .但无论哪种情况 ,Tb3+ 与脱铁乳铁蛋白的结合都不影响蛋白的二级结构 .

The binding of Tb 3+ and Eu 3+ to apolactoferrin has been studied on the basis of the changes in differential UV, fluorescence and CD spectra. It was found that Tb 3+ and Eu 3+ are able to bind to two ferric binding sites and the corresponding conditional equilibrium constants for the Tb 3+ binding were determined to be lg K 1=8.48±0.24, lg K 2=6.72±0.18(0.10 mol/L NaCl, 0.10 mol/L Hepes, pH 7.4 and 25 ℃),but Tb 3+ can not displace the ferric ions from lactoferrin. By lower Tb 3+ /Lf ratio, the conformation became more compact with tryptophane residues exposed to more hydrophobic environment. When more Tb 3+ bound to Lf , the conformation changed in other direction. Anyhow, Tb 3+ binding did not affect the secondary structure.