Fourier transform infrared difference spectroscopy and fluorescence spectroscopic techniques have been used to obtain information about substrate-induced structural changes of the melibiose permease mutant R149C, compared with the Cys-less, which were reconstituted into liposomes. ATR-FTIR evidences show that Na+-induced difference spectra of R149C and Cys-less are similar. However, Na+ induces some new peaks for R149C mutant permease. This means that replacement of Arg-149 by Cys may affect the structure of MelB, and then affect the binding of Na+. Melibiose-induced difference spectra of R149C in the presence of Na+ show some peaks in the amide I region not seen in Cys-less, corresponding to turns, β-sheets, α-helix changes. This suggests that R149C mutant permease undergo some different secondary structure changes compared to Cys-less mutant permease, when binding melibiose. Comparison of the permease intrinsic fluorescence variations of R149C and Cys-less indicate that there are similar substrate binding properties between R149C and Cys-less. When analyzing the effects of different sugars it appears that the R149C mutant is more sensitive to the sugar. All these data indicate that replacement of Arg-149 by Cys will affect Na+ and sugar binding, and enhance the selectivity and sensitivity to sugars.

中文翻译：

---

通过诱变Arg149改变底物诱导的大肠杆菌半乳糖糖通透酶的构象变化

与重构为脂质体的半胱氨酸相比，已经使用傅里叶变换红外差光谱法和荧光光谱技术获得了关于半乳糖通透酶突变体R149C的底物诱导的结构变化的信息。ATR-FTIR证据表明Na +诱导的R149C和Cys-less的差异光谱相似。但是，Na +会诱导R149C突变体通透酶出现一些新峰。这意味着用Cys取代Arg-149可能会影响MelB的结构，然后影响Na +的结合。在Na +存在下，Me149诱导的R149C差异光谱显示，在酰胺I区域中出现了一些在Cys-less少的峰，这对应于匝数，β-折叠，α-螺旋变化。这表明，与无半胱氨酸的突变体通透酶相比，R149C突变体通透酶在结合黑松糖时会发生一些不同的二级结构变化。比较R149C和Cys-less的通透酶固有荧光变化表明，R149C和Cys-less之间具有相似的底物结合特性。分析不同糖的作用时，似乎R149C突变体对糖更敏感。所有这些数据表明，用Cys取代Arg-149将影响Na +和糖的结合，并增强对糖的选择性和敏感性。分析不同糖的作用时，似乎R149C突变体对糖更敏感。所有这些数据表明，用Cys取代Arg-149将影响Na +和糖的结合，并增强对糖的选择性和敏感性。分析不同糖的作用时，似乎R149C突变体对糖更敏感。所有这些数据表明，用Cys取代Arg-149将影响Na +和糖的结合，并增强对糖的选择性和敏感性。