同步荧光光谱探究头孢西丁钠与溶菌酶的结合机制

在模拟生理学条件下(pH=7.40),采用同步荧光法研究了头孢西丁钠(CFXS)和溶菌酶(LYSO)中的荧光基团酪氨酸(Tyr)残基、色氨酸(Trp)残基之间的相互作用。结果表明:CFXS以静态猝灭的方式猝灭LYSO中的Tyr、Trp残基的荧光,结合位点数n≈1。310 K时,Tyr与Trp残基反应的荧光猝灭比率分数NSFQR(Trp)(60.25%)>NSFQR(Tyr)(39.75%),结合位置更靠近Trp残基。Hill系数nH约为1,表明CFXS与LYSO中Tyr与Trp残基的结合不会影响后继配体与蛋白质的结合。CFXS与LYSO中Tyr残基的药物结合率W(Q)为0.19%~0.13%,Trp残基的药物结合率W(Q)为0.23%~0.14%,游离的药物含量几乎不变,这表明CFXS与LYSO中Tyr与Trp残基的结合基本不影响药物的疗效。Tyr残基的蛋白结合率W(B)为52.69%~54.67%,Trp残基的蛋白结合率W(B)为67.67%~69.39%,因此,蛋白中游离的氨基酸残基数目会明显降低。CFXS-LYSO结合体系的主要作用力类型是疏水作用,分子对接结果表明CFXS与LYSO之间还存在氢键作用,且两者的最佳结合位置在LYSO的活性中心附近,两者的结合改变了活性中心处氨基酸残基的微环境。

Study on the binding mechanism of cefoxitin sodium to lysozyme by synchronous fluorescence spectroscopy

Under simulated physiological conditions(pH=7.40),the interaction between tyrosine(Tyr)residue and tryptophan(Trp)residue in lysozyme(LYSO)and cefoxitin sodium(CFXS)was studied using synchronous fluorescence spectroscopy.The results showed that CFXS quenched the fluorescence of Tyr and Trp residue in LYSO by static quenching,and that the number of binding sites n was nearly 1.At 310 K,the fluorescence quenching ratio of CFXS with Trp residue N SFQR(Trp)(60.25%)was higher than that of N SFQR(Tyr)(39.75%),indicating that the binding position was closer to the Trp residue.The Hill coefficient n H was about 1,indicating that the binding of CFXS to the Tyr and Trp residues in LYSO did not affect the binding of subsequent ligands to proteins.The drug binding rate of CFXS to Tyr residue in LYSO was 0.19%to 0.13%,and the drug′s binding rate to Trp residue was 0.23%to 0.14%,respectively.The content of the free drug was almost unchanged.The results showed that the combination of Tyr and Trp residue in LYSO and CFXS did not affect the efficacy of the drug.The protein binding rate of Tyr residue was 52.69%to 54.67%,and the protein binding rate of Trp residue was 67.67%to 69.39%,implying the amount of free amino acid residue in the protein decreased significantly.The main force of the CFXS-LYSO binding system was a hydrophobic interaction.The results of molecular docking showed that there was still a hydrogen bond between the CFXS and LYSO,and the best binding position was near to the active center of the LYSO.The combination of the two substances changed the microenvironment for the amino acid residue at the active center.