Lopatniuk Mariia, Myronovskyi Maksym, Luzhetskyy Andriy
Department of Pharmaceutical Biotechnology, Saarland University , 66123 Saarbrücken, Germany.
Helmholtz-Institute for Pharmaceutical Research , Saarland Campus, Building C2.3, 66123 Saarbrücken, Germany.
ACS Chem Biol. 2017 Sep 15;12(9):2362-2370. doi: 10.1021/acschembio.7b00359. Epub 2017 Aug 16.
The incorporation of noncanonical amino acids (ncAAs) with different side chains into a peptide is a promising technique for changing the functional properties of that peptide. Of particular interest is the incorporation of ncAAs into peptide-derived natural products to optimize their biophysical properties for medical and industrial applications. Here, we present the first instance of ncAA incorporation into the natural product cinnamycin in streptomycetes using the orthogonal pyrrolysyl-tRNA synthetase/tRNA pair from Methanosarcina barkeri. This approach allows site-specific incorporation of ncAAs via the read-through of a stop codon by the suppressor tRNA, which can carry different pyrrolysine analogues. Five new deoxycinnamycin derivatives were obtained with three distinct pyrrolysine analogues incorporated into diverse positions of the antibiotic. The combination of partial hydrolysis and MS/MS fragmentation analysis was used to verify the exact position of the incorporation events. The introduction of ncAAs into different positions of the peptide had opposite effects on the peptide's biological activity.
将具有不同侧链的非标准氨基酸（ncAAs）掺入肽中是一种改变该肽功能特性的很有前景的技术。特别令人感兴趣的是将ncAAs掺入肽衍生的天然产物中，以优化其用于医学和工业应用的生物物理性质。在此，我们展示了使用来自巴氏甲烷八叠球菌的正交吡咯赖氨酸 - tRNA合成酶/tRNA对，首次在链霉菌中将ncAAs掺入天然产物肉桂霉素的实例。这种方法允许通过抑制性tRNA对终止密码子的通读进行ncAAs的位点特异性掺入，该抑制性tRNA可携带不同的吡咯赖氨酸类似物。获得了五种新的脱氧肉桂霉素衍生物，其中三种不同的吡咯赖氨酸类似物被掺入到抗生素的不同位置。使用部分水解和MS/MS碎片分析相结合的方法来验证掺入事件的确切位置。将ncAAs引入肽的不同位置对肽的生物活性产生了相反的影响。
