The Role of a Nonribosomal Peptide Synthetase in l-Lysine Lactamization During Capuramycin Biosynthesis.
The Role of a Nonribosomal Peptide Synthetase in l-Lysine Lactamization During Capuramycin Biosynthesis.
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DOI：
10.1002/cbic.201500701
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发表时间：
2016-05-03
期刊：
影响因子：
--
通讯作者：
Van Lanen SG
中科院分区：
文献类型：
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作者：
Liu X;Jin Y;Cui Z;Nonaka K;Baba S;Funabashi M;Yang Z;Van Lanen SG
辣椒霉素是几种已知的含有 l-Lys 衍生的 l-α-氨基-ε-己内酰胺 (l-ACL) 的天然产物之一。辣椒霉素中 l-ACL 的 α-氨基通过酰胺键与不饱和己糖醛酸成分相连，此前已证明该酰胺键源自不依赖于 ATP 的酶促途径。使用体内和体外相结合的方法，预测的三域非核糖体肽合成酶 CapU 在功能上被表征为 ATP 依赖性酰胺键形成催化剂，负责 l-ACL 中发现的剩余酰胺键的生物合成。结果与 CapU 的腺苷酸化结构域作为 L-Lys 活化和相邻硫醇化结构域硫酯化的基本催化成分一致。然而，与预期相反，内酰胺化不需要任何额外的结构域或蛋白质，并且可能是非酶促事件。这些结果为研究是否在其他含有 l-ACL 的天然产物的生物合成中采用了类似的 NRPS 介导的机制奠定了基础，同样有趣的是，如何在众多含有未修饰的 l-Lys 的 NRPS 衍生肽中避免自发内酰胺化。使用体内和体外方法描述了作为辣椒霉素抗分枝杆菌抗生素成分的氨基己内酰胺的生物合成机制。该过程由 L-Lys 的腺苷酸化和硫酯化引发，并通过明显的非酶分子内氨解来强调。
Capuramycins are one of several known natural products that contain an l-Lys-derived l-α-amino-ε-caprolactam (l-ACL). The α-amino group of l-ACL in capuramycins is linked to an unsaturated hexuronic acid component through an amide bond that was previously shown to originate via an ATP-independent enzymatic route. Using a combined in vivo and in vitro approach, a predicted tridomain nonribosomal peptide synthetase CapU is functionally characterized here as the ATP-dependent amide bond-forming catalyst responsible for the biosynthesis of the remaining amide bond found in l-ACL. The results are consistent with the adenylation domain of CapU as the essential catalytic component for l-Lys activation and thioesterification of the adjacent thiolation domain. However, in contrast to expectations, lactamization does not require any addi-tional domains or proteins and is likely a nonenzymatic event. The results set the stage to examine whether a similar NRPS-mediated mechanism is employed in the biosynthesis of other l-ACL-containing natural products and, just as intriguing, how spontaneous lactamization is avoided in the numerous NRPS-derived peptides that contain an unmodified l-Lys. The biosynthetic mechanism for the aminocaprolactam that is a component of the capuramycin antimycobacterial antibiotics was delineated using an in vivo and in vitro approach. The process is initiated by adenylation and thioesterification of l-Lys and highlighted by an apparently nonenzymatic intramolecular aminolysis.