Thoppil Anu A, Sharma Ruchika, Kishore Nand
Department of Chemistry, Indian Institute of Technology Bombay, Powai, Mumbai 400076, India.
Biopolymers. 2008 Oct;89(10):831-40. doi: 10.1002/bip.21021.
Binding of the antibiotic drug carbenicillin to bovine serum albumin (BSA) has been studied using isothermal titration calorimetry (ITC) in combination with fluorescence and circular dichroism (CD) spectroscopies. The thermodynamic parameters of binding have been evaluated as a function of temperature, ionic strength, and in the presence of anionic, cationic and nonionic surfactants, tetrabutylammonium bromide, and sucrose. The values of van't Hoff enthalpy do not agree with the calorimetric enthalpy indicating conformational changes in the protein upon drug binding. These observations are supported by the intrinsic fluorescence and CD spectroscopic measurements. A reduction in the binding affinity of carbenicillin to BSA is observed with increase in ionic strength of the solution, thereby suggesting, prevailing of electrostatic interactions in the binding process. The involvement of hydrophobic interactions in the binding of the drug to the protein is also indicated by a slight reduction in binding constant in the presence of tetrabutylammonium bromide. The experiments in the presence of sucrose suggest that hydrogen bonding is perhaps not dominant in the binding. The anionic surfactant sodium dodecyl sulphate (SDS) is observed to completely interfere in the ionic interactions in addition to its partial denaturing capacity. However, the presence of cationic surfactant hexadecyl trimethylammonium bromide (HTAB) and nonionic surfactant Triton-X 100 induce a slight reduction in the values of binding affinity. These calorimetric and spectroscopic results, provide quantitative information on the binding of carbenicillin to BSA and suggests that the binding is dominated by electrostatic interactions with contribution from hydrophobic interactions.
已使用等温滴定量热法（ITC）结合荧光光谱和圆二色性（CD）光谱研究了抗生素药物羧苄青霉素与牛血清白蛋白（BSA）的结合。结合的热力学参数已根据温度、离子强度以及在阴离子、阳离子和非离子表面活性剂、四丁基溴化铵和蔗糖存在的情况下进行了评估。范特霍夫焓值与量热焓值不一致，表明药物结合后蛋白质发生了构象变化。这些观察结果得到了固有荧光和CD光谱测量的支持。随着溶液离子强度的增加，观察到羧苄青霉素与BSA的结合亲和力降低，从而表明在结合过程中静电相互作用占主导。在四丁基溴化铵存在下结合常数略有降低也表明疏水相互作用参与了药物与蛋白质的结合。在蔗糖存在下的实验表明氢键在结合中可能不占主导。观察到阴离子表面活性剂十二烷基硫酸钠（SDS）除了具有部分变性能力外，还完全干扰离子相互作用。然而，阳离子表面活性剂十六烷基三甲基溴化铵（HTAB）和非离子表面活性剂 Triton-X 100 的存在会导致结合亲和力值略有降低。这些量热和光谱结果提供了关于羧苄青霉素与BSA结合的定量信息，并表明结合主要由静电相互作用主导，疏水相互作用也有贡献。
