Xu Meimei, Wilderman P Ross, Peters Reuben J
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, USA.
Proc Natl Acad Sci U S A. 2007 May 1;104(18):7397-401. doi: 10.1073/pnas.0611454104. Epub 2007 Apr 24.
There have been few insights into the biochemical origins of natural product biosynthesis from primary metabolism. Of particular interest are terpene synthases, which often mediate the committed step in particular biosynthetic pathways so that alteration of their product outcome is a key step in the derivation of novel natural products. These enzymes also catalyze complex reactions of significant mechanistic interest. Following an evolutionary lead from two recently diverged, functionally distinct diterpene synthase orthologs from different subspecies of rice, we have identified a single residue that can switch product outcome. Specifically, the mutation of a conserved isoleucine to threonine that acts to convert not only the originally targeted isokaurene synthase into a specific pimaradiene synthase but also has a much broader effect, which includes conversion of the ent-kaurene synthases found in all higher plants for gibberellin phytohormone biosynthesis to the production of pimaradiene. This surprisingly facile switch for diterpene synthase catalytic specificity indicates the ease with which primary (gibberellin) metabolism can be subverted to secondary biosynthesis and may underlie the widespread occurrence of pimaradiene-derived natural products. In addition, because this isoleucine is required for the mechanistically more complex cyclization to tetracyclic kaurene, whereas substitution with threonine "short-circuits" this mechanism to produce the "simpler" tricyclic pimaradiene, our results have some implications regarding the means by which terpene synthases specify product outcome.
关于天然产物生物合成从初级代谢的生化起源，人们了解甚少。特别值得关注的是萜烯合酶，它们常常介导特定生物合成途径中的关键步骤，因此改变其产物结果是新型天然产物衍生过程中的关键一步。这些酶还催化具有重要机制研究价值的复杂反应。沿着来自水稻不同亚种的两个最近分化、功能不同的二萜合酶直系同源物的进化线索，我们鉴定出了一个能够改变产物结果的单一残基。具体而言，将一个保守的异亮氨酸突变为苏氨酸，这不仅能将原本靶向的异贝壳杉烯合酶转化为特定的海松二烯合酶，而且具有更广泛的影响，包括将所有高等植物中用于赤霉素植物激素生物合成的内贝壳杉烯合酶转化为海松二烯的产生。二萜合酶催化特异性这种出人意料的简单转变表明，初级（赤霉素）代谢可以很容易地转向次级生物合成，这可能是海松二烯衍生的天然产物广泛存在的基础。此外，由于这个异亮氨酸对于机制上更复杂的四环贝壳杉烯环化是必需的，而用苏氨酸替代会“短路”该机制以产生“更简单”的三环海松二烯，我们的结果对于萜烯合酶确定产物结果的方式具有一定启示。
