Rising Kathleen A, Crenshaw Charisse M, Koo Hyun Jo, Subramanian Thangaiah, Chehade Kareem A H, Starks Courtney, Allen Keith D, Andres Douglas A, Spielmann H Peter, Noel Joseph P, Chappell Joe
†Departments of Pharmaceutical Sciences, ‡Cellular and Molecular Biochemistry, §Chemistry, ⊥Center for Structural Biology, ∥Markey Cancer Center, University of Kentucky, Lexington, Lexington, Kentucky, United States.
∇Howard Hughes Medical Institute, Salk Institute, La Jolla, California 92037, United States.
ACS Chem Biol. 2015 Jul 17;10(7):1729-36. doi: 10.1021/acschembio.5b00145. Epub 2015 May 4.
As part of an effort to identify substrate analogs suitable for helping to resolve structural features important for terpene synthases, the inhibition of 5-epi-aristolochene biosynthesis from farnesyl diphosphate (FPP) by the tobacco 5-epi-aristolochene synthase incubated with anilinogeranyl diphosphate (AGPP) was examined. The apparent noncompetitive nature of the inhibition supported further assessment of how AGPP might be bound to crystallographic forms of the enzyme. Surprisingly, the bound form of the inhibitor appeared to have undergone a cyclization event consistent with the native mechanism associated with FPP catalysis. Biocatalytic formation of a novel 13-membered macrocyclic paracyclophane alkaloid was confirmed by high-resolution GC-MS and NMR analysis. This work provides insights into new biosynthetic means for generating novel, functionally diversified, medium-sized terpene alkaloids.
作为鉴定适合帮助解析对萜类合酶重要的结构特征的底物类似物工作的一部分，研究了与苯胺基香叶基二磷酸（AGPP）一起孵育的烟草5-表-马兜铃烯合酶对法呢基二磷酸（FPP）生物合成5-表-马兜铃烯的抑制作用。抑制作用的明显非竞争性性质支持了对AGPP如何与该酶的晶体形式结合的进一步评估。令人惊讶的是，抑制剂的结合形式似乎经历了与FPP催化相关的天然机制一致的环化事件。通过高分辨率气相色谱-质谱联用（GC-MS）和核磁共振（NMR）分析证实了一种新型13元大环对环芳烷生物碱的生物催化形成。这项工作为生成新型、功能多样的中等大小萜类生物碱提供了新的生物合成方法的见解。
