Abstract
Terpenes (isoprenoids), derived from isoprenyl pyrophosphates, are versatile natural compounds that act as metabolism mediators, plant volatiles, and ecological communicators. Divergent evolution of homomeric prenyltransferases (PTSs) has allowed PTSs to optimize their active-site pockets to achieve catalytic fidelity and diversity. Little is known about heteromeric PTSs, particularly the mechanisms regulating formation of specific products. Here, we report the crystal structure of the (LSU . SSU)(2)-type (LSU/SSU = large/small subunit) heterotetrameric geranyl pyrophosphate synthase (GPPS) from mint (Mentha piperita). The LSU and SSU of mint GPPS are responsible for catalysis and regulation, respectively, and this SSU lacks the essential catalytic amino acid residues found in LSU and other PTSs. Whereas no activity was detected for individually expressed LSU or SSU, the intact (LSU . SSU)(2) tetramer produced not only C(10)-GPP at the beginning of the reaction but also C(20)-GGPP (geranylgeranyl pyrophosphate) at longer reaction times. The activity for synthesizing C(10)-GPP and C(20)-GGPP, but not C(15)-farnesyl pyrophosphate, reflects a conserved active-site structure of the LSU and the closely related mustard (Sinapis alba) homodimeric GGPPS. Furthermore, using a genetic complementation system, we showed that no C(20)-GGPP is produced by the mint GPPS in vivo. Presumably through protein-protein interactions, the SSU remodels the active-site cavity of LSU for synthesizing C(10)-GPP, the precursor of volatile C(10)-monoterpenes.
Highlights
Linear prenyl pyrophosphates (LPPs) are the precursors for the more than 55,000 terpenes that have been identified in various organisms
X-ray crystallographic analysis of the Mp geranyl pyrophosphate synthase (GPPS) revealed a novel (LSU·SSU)2 architecture composed of two LSU·SSU heterodimers (Figure 2)
Our structural and mutagenetic studies offer a new insight into how the product specificity and fidelity of the (LSU.SSU)2-type M. piperita GPPS (Mp GPPS) is determined via intersubunit regulation through a novel molecular mechanism
Summary
Linear prenyl pyrophosphates (LPPs) are the precursors for the more than 55,000 terpenes (isoprenoids) that have been identified in various organisms. Many are essential for important biological processes, such as protein prenylation (Ras, Rab, and Rho), proper functioning of the electron transport chain (quinine and heme a), glycoprotein biosynthesis (dolichol), and the metabolism of growth hormones (gibberellin, cytokinin, and sterol) (Liang et al, 2002; McTaggart, 2006; Gershenzon and Dudareva, 2007; Christianson, 2008; Kirby and Keasling, 2009). Some terpenes have considerable commercial interest as medicines (taxol for anticancer and artemisinin for antimalaria), flavors and fragrances (menthol and linalool), and nutritional supplements (carotenoid and vitamin A) (Pichersky et al, 2006; Gershenzon and Dudareva, 2007; Kirby and Keasling, 2009).
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