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用荧光光谱、等温滴定微量热(ITC)及分子模建3种方法研究了人尿胰蛋白酶抑制剂(UTI)与1-苯胺基-8-萘磺酸(ANS)的相互作用.结果表明:在UTI上有4个以静电相互作用为主要作用力的特异性的ANS结合位点,分别命名为位点Ⅰ,Ⅱ,Ⅲ和Ⅳ;其中位点Ⅰ位于UTI上结构域Ⅱ的第98位色氨酸(Trp98)附近,位点Ⅱ位于结构域Ⅰ和结构域Ⅱ的相互作用区,位点Ⅲ与位点Ⅳ位于结构域Ⅰ;这4个特异性结合位点所处的区域疏水性较强.ITC实验测得另外5个非特异性结合位点,其主要相互作用力是以ANS的磺酸基与UTI分子表面的带正电的基团间形成的盐键,表明在中性缓冲液中,有5个带正电的氨基酸残基暴露在UTI分子表面.
The interaction between human urinary trypsin inhibitor (UTI) and 1-anilino-8-naphthalenesulfonic acid (ANS) was studied by fluorescence spectroscopy, isothermal titration calorimetry (ITC) and molecular modeling. There are four specific ANS binding sites on the UTI that are mainly interacting with electrostatic interactions and are named as sites I, II, III and IV, respectively. The site I is located at position 98 of domain II of UTI In the vicinity of tryptophan (Trp98), the site Ⅱ is located in the interaction domain of domain Ⅰ and domain Ⅱ, the site Ⅲ and site Ⅳ are located in domain I. The region where these 4 specific binding sites are located is hydrophobic Stronger.ITC experiment measured the other five non-specific binding sites, the main interaction between the ANS sulfonic acid group and the UTI molecule surface positively charged groups formed between the salt bond, indicating that in the neutral buffer Liquid, there are five positive amino acid residues exposed on the surface of UTI molecules.