Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
Institute of Biostructures and Bioimaging, CNR, Naples, Italy.
Protein Sci. 2024 Sep;33(9):e5146. doi: 10.1002/pro.5146.
D2 is a structural and cooperative domain of Thermotoga maritima Arginine Binding Protein, that possesses a remarkable conformational stability, with a denaturation temperature of 102.6°C, at pH 7.4. The addition of potassium thiocyanate causes a significant decrease in the D2 denaturation temperature. The interactions of thiocyanate ions with D2 have been studied by means of isothermal titration calorimetry measurements and molecular dynamics simulations. It emerged that: (a) 20-30 thiocyanate ions interact with the D2 surface and are present in its first solvation shell; (b) each of them makes several contacts with protein groups, both polar and nonpolar ones. The addition of guanidinium thiocyanate causes a marked destabilization of the D2 native state, because both the ions are denaturing agents. However, on adding to the solution containing D2 and guanidinium thiocyanate a stabilizing agent, such as TMAO, sucrose or sodium sulfate, a significant increase in denaturation temperature occurs. The present results confirm that counteraction is a general phenomenon for globular proteins.
D2 是海洋栖热菌精氨酸结合蛋白的结构和协同域，具有显著的构象稳定性，其变性温度为 102.6°C，在 pH 值为 7.4 时。添加硫氰酸钾会导致 D2 变性温度显著降低。通过等温热滴定法测量和分子动力学模拟研究了硫氰酸根离子与 D2 的相互作用。结果表明：（a）20-30 个硫氰酸根离子与 D2 表面相互作用，并存在于其第一个溶剂化壳中；（b）它们中的每一个都与蛋白质基团，包括极性和非极性基团，形成多个接触。添加胍基硫氰酸盐会显著破坏 D2 的天然状态，因为这两种离子都是变性剂。然而，在向含有 D2 和胍基硫氰酸盐的溶液中添加稳定剂，如 TMAO、蔗糖或硫酸钠，变性温度会显著升高。本研究结果证实，拮抗作用是球状蛋白质的普遍现象。
