Gadow A, Vater J, Schlumbohm W, Palacz Z, Salnikow J, Kleinkauf H
Eur J Biochem. 1983 May 2;132(2):229-34. doi: 10.1111/j.1432-1033.1983.tb07352.x.
The reactive thioester complexes of gramicidin S synthetase with substrate amino acids and intermediate peptides are slowly hydrolyzed in neutral buffer solutions under mild conditions. Fully active enzyme is recovered. These processes are strongly accelerated by certain thiol protective agents. In the presence of 1 mM dithioerythritol the half-life times of these hydrolysis reactions are in the range of 1-90 h at 3 degrees C. The thioester complex of gramicidin S synthetase 2 (GS2, the heavy enzyme) with the tripeptide DPhe-Pro-Val is distinguished by the highest stability of all these intermediates. A different decomposition pattern is observed for the thioester complex of GS2 with LOrn. Here 3-amino-2-piperidone (cyclo-LOrn) is formed in a rapid cyclization reaction. This product specifically blocks the activation center of GS2 for LOrn at the thioester binding site. All other activation reactions of gramicidin S synthetase are unaffected. A procedure for a specific labelling of the reaction centers of the multienzyme is outlined.
短杆菌肽S合成酶与底物氨基酸和中间肽形成的反应性硫酯复合物在温和条件下于中性缓冲溶液中缓慢水解。可回收完全活性的酶。某些硫醇保护剂能强烈加速这些过程。在1 mM二硫苏糖醇存在下，这些水解反应在3℃时的半衰期为1至90小时。短杆菌肽S合成酶2（GS2，重酶）与三肽DPhe-Pro-Val形成的硫酯复合物在所有这些中间体中稳定性最高。观察到GS2与LOrn形成的硫酯复合物有不同的分解模式。在此，3-氨基-2-哌啶酮（环-LOrn）在快速环化反应中形成。该产物在硫酯结合位点特异性阻断GS2对LOrn的激活中心。短杆菌肽S合成酶的所有其他激活反应均不受影响。概述了一种对多酶反应中心进行特异性标记的方法。
