Site-Directed and Global Incorporation of Orthogonal and Isostructural Noncanonical Amino Acids into the Ribosomal Lasso Peptide Capistruin
Site-Directed and Global Incorporation of Orthogonal and Isostructural Noncanonical Amino Acids into the Ribosomal Lasso Peptide Capistruin
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DOI：
10.1002/cbic.201402558
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发表时间：
2015-02-09
期刊：
影响因子：
3.2
通讯作者：
Suessmuth, Roderich D.
中科院分区：
文献类型：
--
作者：
Al Toma, Rashed S.;Kuthning, Anja;Suessmuth, Roderich D.
通过两种不同的方法进行肽抗生素的结构多样性的扩展。基于补充的掺入（SPI）和终止密码子抑制（SCS）方法用于将同构和正交非规范氨基酸（ncAA）共翻译掺入到套索肽capistruin中。SPI方法采用两种ncAA，SCS方法采用五种ncAA;每种ncAA均探测ncAA在分子战略位置的掺入。通过HR-ESI-MS对组装的评估证明SCS方法更成功。生物正交化学用于含有ncAA Alk（Ne-Alloc-l-赖氨酸）代替Ala的capistruin同源物Cap_Alk10的生物合成后修饰。第二代Hoveyda-Grubbs催化剂用于与Cap_Alk10和烯丙醇的体外复分解反应，这为生物合成后的修饰提供了选择。合成生物学的使用允许从扩展的氨基酸库体内生产新的基于肽的抗生素。
Expansion of the structural diversity of peptide antibiotics was performed through two different methods. Supplementation-based incorporation (SPI) and stop-codon suppression (SCS) approaches were used for co-translational incorporation of isostructural and orthogonal noncanonical amino acids (ncAAs) into the lasso peptide capistruin. Two ncAAs were employed for the SPI method and five for the SCS method; each of them probing the incorporation of ncAAs in strategic positions of the molecule. Evaluation of the assembly by HR-ESI-MS proved more successful for the SCS method. Bio-orthogonal chemistry was used for post-biosynthetic modification of capistruin congener Cap_Alk10 containing the ncAA Alk (Ne-Alloc-l-lysine) instead of Ala. A second-generation Hoveyda-Grubbs catalyst was used for an in vitro metathesis reaction with Cap_Alk10 and an allyl alcohol, which offers options for post-biosynthetic modifications. The use of synthetic biology allows for the in vivo production of new peptide-based antibiotics from an expanded amino acid repertoire.