等温滴定量热法研究核糖开关aac与氨基糖苷类抗生素的相互作用  

Isothermal Titration Calorimetry Studies on the Interaction of Riboswitch aac with Aminoglycosides

作　　者：石会刚 陈东戎[1] 

机构地区：[1]复旦大学生物医学研究院,上海200032

出　　处：《生物物理学报》2015年第3期241-250,共10页Acta Biophysica Sinica

基　　金：国家自然科学基金重点项目(31330022)~~

摘　　要：核糖开关(riboswitch)是位于m RNA非编码区域的RNA元件,可与小分子配体结合导致结构转变,进而调控下游基因的表达。核糖开关aac能够特异性结合氨基糖苷类抗生素,从而调控氨基糖苷类抗生素抗性基因的表达。目前,核糖开关aac与氨基糖苷类抗生素相互作用的位点和机制尚不清楚。作者利用等温滴定量热法对核糖开关aac与氨基糖苷类抗生素的结合亲和力及结合热力学性质进行了研究,并初步探讨了点突变对相互作用的影响。结果发现,西索米星、庆大霉素、G418、奈替霉素和巴龙霉素能够特异性结合核糖开关aac,而阿米卡星、卡那霉素A、链霉素、链丝菌素、新霉胺、新霉素及核糖霉素与核糖开关aac无特异性相互作用;核糖开关aac与氨基糖苷类抗生素的结合热力学性质表明二者通过氢键和范德华力发生相互作用。西索米星滴定核糖开关aac突变体的结果表明,U68和A18位点很可能与氨基糖苷类抗生素形成氢键,A13和A44位点对于相互作用也有影响。这些结果对进一步研究核糖开关aac与氨基糖苷类抗生素相互作用的位点和机制具有重要的参考价值。Riboswitch is a regulatory element located at mRNA untranslated regions, and it can bind with small molecules, regulating gene expression. Riboswitch aac is a novel riboswitch which can bind with aminoglycosides, and regulates aminoglycoside resistance gene expression. However, the mechanism of riboswitch-aminoglycosides interaction is still unknown. In this paper, the authors investigated the binding affinity and thermodynamics of riboswitch-aminoglycosides interactions by isothermal titration calorimetry （ITC） and the potential influence of point mutations on the interactions were also studied. The ITC data reveals that sisomicin, gentamicin, G418, netilmicin and paromomycin bind riboswitch aac with high affinity, while other aminoglycosides such as amikacin, kanamycin A, neomycin, neamine, streptomycin, streptothricin and ribostamycin don＇t bind riboswitch aac. The thermodynamics of riboswitch-aminoglycosides interaction reveals that the interaction is driven by enthalpy change, and the enthalpy change provides important insights into the contribution of hydrogen bond and Van der Waals＇ force. They also found that U68 and A18 of riboswitch aac are potential binding sites and A13, A44 are also essential for riboswitch-aminoglycosides interaction. This study provides important reference for further study on mechanisms of riboswitches- aminoglycosides.

关 键 词：核糖开关 氨基糖苷类抗生素 等温滴定量热法 热力学 

分 类 号：Q522[生物学—生物化学]