16-Aza-ent-beyerane and 16-Aza-ent-trachylobane: Potent Mechanism-Based Inhibitors of Recombinant ent-Kaurene Synthase from Arabidopsis thaliana
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- 作者：Arnab Roy ; Frank G. Roberts ; P. Ross Wilderman ; Ke Zhou ; Reuben J. Peters ; Robert M. Coates
- 刊名：Journal of the American Chemical Society
- 出版年：2007
- 出版时间：October 17, 2007
- 年：2007
- 卷：129
- 期：41
- 页码：12453 - 12460
- 全文大小：153K
- 年卷期：v.129,no.41(October 17, 2007)
- ISSN：1520-5126
文摘
The secondary ent-beyeran-16-yl carbocation (7) is a key branch point intermediate in mechanisticschemes to rationalize the cyclic structures of many tetra- and pentacyclic diterpenes, including ent-beyerene,ent-kaurene, ent-trachylobane, and ent-atiserene, presumed precursors to >1000 known diterpenes. Toevaluate these mechanistic hypotheses, we synthesized the heterocyclic analogues 16-aza-ent-beyerane(12) and 16-aza-ent-trachylobane (13) by means of Hg(II)- and Pb(IV)-induced cyclizations onto the 12double bonds of tricyclic intermediates bearing carbamoylmethyl and aminomethyl groups at C-8. The13,16-seco-16-norcarbamate (20a) was obtained from ent-beyeran-16-one oxime (17) by Beckmannfragmentation, hydrolysis, and Curtius rearrangement. The aza analogues inhibited recombinant ent-kaurenesynthase from Arabidopsis thaliana (GST-rAtKS) with inhibition constants (IC50 = 1 × 10-7 and 1 × 10-6M) similar in magnitude to the pseudo-binding constant of the bicyclic ent-copalyl diphosphate substrate(Km = 3 × 10-7 M). Large enhancements of binding affinities (IC50 = 4 × 10-9 and 2 × 10-8 M) wereobserved in the presence of 1 mM pyrophosphate, which is consistent with a tightly bound ent-beyeranyl+/pyrophosphate- ion pair intermediate in the cyclization-rearrangement catalyzed by this diterpene synthase.The weak inhibition (IC50 = 1 × 10-5 M) exhibited by ent-beyeran-16-exo-yl diphosphate (11) and its failureto undergo bridge rearrangement to kaurene appear to rule out the covalent diphosphate as a freeintermediate. 16-Aza-ent-beyerane is proposed as an effective mimic for the ent-beyeran-16-yl carbocationwith potential applications as an active site probe for the various ent-diterpene cyclases and as a novel,selective inhibitor of gibberellin biosynthesis in plants.