Enhanced macrocyclizing activity of the thioesterase from tyrocidine synthetase in presence of nonionic detergent.
Enhanced macrocyclizing activity of the thioesterase from tyrocidine synthetase in presence of nonionic detergent.
复制标题
在非离子去污剂存在下，酪氨酸合成酶的硫酯酶的大环化活性增强。
DOI：
10.1016/j.chembiol.2004.09.003
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Walsh,ChristopherT
中科院分区：
文献类型：
--
作者：
Yeh,Ellen;Lin,Hening;Clugston,SusanL;Kohli,RahulM;Walsh,ChristopherT
由多模块非核糖体肽合成酶（NRPS）硫酯酶结构域进行的大环化是许多生物活性肽生物合成的关键步骤。从酪氨酸合成酶切下的硫酯酶是一种多用途的大环化催化剂，并且是化学酶促合成多种环肽的有用工具。然而，它的实用性是有限的，其催化活性的寿命短，以及显着的流量的酰基酶中间体水解。在临界胶束浓度以上添加Brij 58（一种非离子去污剂）对酶活性具有显著影响：催化活性延长至>60 min，环化（但不是水解）速率增加6倍，导致环化产物产率净增加150- 300倍。这种增强的活性允许酪肽十肽的固相文库的酶促大环化，以鉴定在Orn 9位置处的可接受的取代，所述取代先前对于多样化是不可接近的。
Macrocyclization carried out by thioesterase domains of multimodular nonribosomal peptide synthetases (NRPSs) is a key step in the biosynthesis of many biologically active peptides. The thioesterase excised from tyrocidine synthetase is a versatile macrocyclization catalyst and a useful tool for chemoenzymatic synthesis of diverse cyclic peptides. However, its utility is limited by its short lifetime of catalytic activity as well as significant flux of the acyl-enzyme intermediate to hydrolysis. The addition of Brij 58, a nonionic detergent, above the critical micelle concentration, has dramatic effects on enzyme activity: catalytic activity is extended to >60 min and the rate of cyclization (but not hydrolysis) increases 6-fold, resulting in a net 150- to 300-fold increase in cyclic product yields. This enhanced activity allowed enzymatic macrocyclization of a solid phase library of tyrocidine decapeptides to identify acceptable substitutions at the Orn9 position which had previously been inaccessible for diversification.