Molecular Genetics Department and Kluyver Centre for Genomics of Industrial Fermentation, Synthetic Biology Centre, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands.
Chembiochem. 2012 Nov 5;13(16):2433-8. doi: 10.1002/cbic.201200437. Epub 2012 Oct 15.
The nisin leader is believed to be crucial for nisin biosynthesis. Here, by using a construct completely lacking the leader peptide, we show that an up to fivefold-dehydrated leaderless prenisin can be obtained, as judged by MALDI-TOF MS, and that some of these species are biologically active, thus suggesting that at least three lanthionine rings are present. Notably, by expressing the leader peptide in trans together with the leaderless prenisin, we were able to increase the dehydration/cyclization efficiency of both NisB and NisC, but still with limited efficiency until the fifth dehydratable residue (Thr13) was processed, thereby enabling three rings to form. This, for the first time, demonstrates that 1) the leader is not absolutely necessary for the dehydration reaction of class I lantibiotics to occur in vivo; 2) the leader acts in trans in vivo; 3) the leader increases the efficiency of modification. Based on previous work and our current study, a model for the interactions of NisB and NisC with prenisin is proposed, in which the leader induces a more active conformation and/or productive complex formation of the biosynthetic machinery, and, when covalently bound, is involved in increasing the efficiency of dehydration to the C-terminal end of the prenisin substrate molecule.
乳链菌肽的前导肽被认为对其生物合成至关重要。在这里，我们通过使用完全缺乏前导肽的构建体，通过 MALDI-TOF MS 判断可以得到多达五倍脱水的无领导肽前乳链菌肽，并且其中一些具有生物活性，因此表明至少存在三个链桥氨酸环。值得注意的是，通过与无领导肽一起表达前导肽，我们能够提高 NisB 和 NisC 的脱水/环化效率，但直到处理第五个可脱水残基（Thr13）时，效率仍然有限，从而使三个环形成。这首次证明了 1）前导肽对于体内发生 I 类类细菌素的脱水反应不是绝对必需的；2）前导肽在体内起反式作用；3）前导肽提高了修饰效率。基于以前的工作和我们目前的研究，提出了 NisB 和 NisC 与前乳链菌肽相互作用的模型，其中前导肽诱导生物合成机制更活跃的构象和/或更有效的复合物形成，并且当共价结合时，参与增加脱水效率到前乳链菌肽底物分子的 C 末端。
