达托霉素作为新的胰蛋白酶激活剂的初探

Study on Daptomycin as a New Trypsin Activator

首次发现达托霉素是一种新的胰蛋白酶激活剂,当胰蛋白酶与达托霉素的物质的量的比在反应时达到34.05时,达托霉素对胰蛋白酶比活力的激活率平均达到32.92%。通过计算机模拟技术对达托霉素与胰蛋白酶以及达托霉素对胰蛋白酶底物复合物分别进行分子对接,并利用等温滴定量热法(Isothermal titration calorimetry,ITC)验证模拟结果。研究发现达托霉素与胰蛋白酶的活性中心位置很靠近,达托霉素链状中的天冬氨酸的R基与酶活性中心的组氨酸-57发生了氢键相互作用,达托霉素使酶和底物复合物结构更加稳定,从而有利于催化作用。ITC结果表明胰蛋白酶上具有1个达托霉素结合位点,解离常数Kd为17.83μM,摩尔结合焓△H为237.9±28.17 cal/mol,摩尔结合熵△S为22.5 cal/mol·deg,这些结果从热力学角度支持了分子对接结果。研究发现达托霉素除具有抗革兰氏阳性耐药性细菌的功能外,还能促进胰蛋白酶比活这一新的功能,是一种新的胰蛋白酶激活剂。

The activity of trypsin was increased 32. 92 % on average when the amount of substance rate between trypsin and daptomycin is 34. 05, which indicated that daptomycin is a new trypsin activator.Daptomycin-trypsin and daptomycin-trypsin-substrate complex were studied by the molecular docking, and isothermal titration calorimetry（ ITC） was used to support the result of molecular docking. It showed that daptomycin was close to the activity center of trypsin,and the R group of daptomycin was interacted with histidine-57 of trypsin,and this histidine-57 located in the activity center of the enzyme. The complex structure between enzyme and substrate is more stable when the daptomycin is presence,which is beneficial to the catalytic effect. The ITC results demonstrated that a daptomycin binding site was located in trypsin,and the dissociation constant Kd is 17. 83 μM,molar combustion enthalpy △H is 237. 9 ± 28. 17 cal/mol,molar binding entropy △S is 22. 5 cal/mol/deg,and all the results support the molecular docking results. It indicated that daptomycin is a new trypsin activator and it will have application in proteomics and other fields.