Dataset Information

Crystal Structure and Potential Head-to-Middle Condensation Function of a Z,Z-Farnesyl Diphosphate Synthase.

ABSTRACT: Plants produce a wide variety of secondary metabolites in response to adverse environmental factors. Z,Z-Farnesyl diphosphate (Z,Z-FPP), synthesized by Z,Z-farnesyl diphosphate synthase (zFPS), supports the formation of phytochemicals in wild tomatoes. Here, the crystal structure of N-terminal truncated zFPS (?zFPS) was determined. Irregular products including lavandulyl diphosphate and an unknown compound were surprisingly found. Apart from the truncated N-terminus as a functional regulator, structure-based analysis and mutagenesis assays revealed a residue H103 in ?zFPS as one of the key elements to this irregular function. A series of substrate-enzyme complex structures were obtained from ?zFPS-H103Y by co-crystallizing with isopentenyl diphosphate, dimethylallyl thiolodiphosphate, or both. Various substrate-binding modes were revealed. The catalytic mechanisms of both the head-to-tail and head-to-middle reactions in ?zFPS were proposed. Functional switch between the two mechanisms in this enzyme and the essential role played by the flexible C-terminus were elucidated as well.

SUBMITTER: Chan YT

PROVIDER: S-EPMC6044691 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Crystal Structure and Potential Head-to-Middle Condensation Function of a Z,Z-Farnesyl Diphosphate Synthase.

Chan Yueh-Te YT Ko Tzu-Ping TP Yao Shan-Hsueh SH Chen Ya-Wen YW Lee Cheng-Chung CC Wang Andrew H-J AH

ACS omega 20170316 3

Plants produce a wide variety of secondary metabolites in response to adverse environmental factors. Z,Z-Farnesyl diphosphate (Z,Z-FPP), synthesized by Z,Z-farnesyl diphosphate synthase (zFPS), supports the formation of phytochemicals in wild tomatoes. Here, the crystal structure of N-terminal truncated zFPS (ΔzFPS) was determined. Irregular products including lavandulyl diphosphate and an unknown compound were surprisingly found. Ap ...[more]

PMID: 30023621

Similar Datasets

Project description:BackgroundIsoprenoids are the most diverse and abundant group of natural products. In Plasmodium falciparum, isoprenoid synthesis proceeds through the methyl erythritol diphosphate pathway and the products are further metabolized by farnesyl diphosphate synthase (FPPS), turning this enzyme into a key branch point of the isoprenoid synthesis. Changes in FPPS activity could alter the flux of isoprenoid compounds downstream of FPPS and, hence, play a central role in the regulation of a number of essential functions in Plasmodium parasites.MethodsThe isolation and cloning of gene PF3D7_18400 was done by amplification from cDNA from mixed stage parasites of P. falciparum. After sequencing, the fragment was subcloned in pGEX2T for recombinant protein expression. To verify if the PF3D7_1128400 gene encodes a functional rPfFPPS protein, its catalytic activity was assessed using the substrate [4-14C] isopentenyl diphosphate and three different allylic substrates: dimethylallyl diphosphate, geranyl diphosphate or farnesyl diphosphate. The reaction products were identified by thin layer chromatography and reverse phase high-performance liquid chromatography. To confirm the product spectrum formed of rPfFPPS, isoprenic compounds were also identified by mass spectrometry. Apparent kinetic constants KM and Vmax for each substrate were determined by Michaelis-Menten; also, inhibition assays were performed using risedronate.ResultsThe expressed protein of P. falciparum FPPS (rPfFPPS) catalyzes the synthesis of farnesyl diphosphate, as well as geranylgeranyl diphosphate, being therefore a bifunctional FPPS/geranylgeranyl diphosphate synthase (GGPPS) enzyme. The apparent KM values for the substrates dimethylallyl diphosphate, geranyl diphosphate and farnesyl diphosphate were, respectively, 68?±?5 ?M, 7.8?±?1.3 ?M and 2.06?±?0.4 ?M. The protein is expressed constitutively in all intra-erythrocytic stages of P. falciparum, demonstrated by using transgenic parasites with a haemagglutinin-tagged version of FPPS. Also, the present data demonstrate that the recombinant protein is inhibited by risedronate.ConclusionsThe rPfFPPS is a bifunctional FPPS/GGPPS enzyme and the structure of products FOH and GGOH were confirmed mass spectrometry. Plasmodial FPPS represents a potential target for the rational design of chemotherapeutic agents to treat malaria.

Dataset Information

Crystal Structure and Potential Head-to-Middle Condensation Function of a Z,Z-Farnesyl Diphosphate Synthase.

Publications

Crystal Structure and Potential Head-to-Middle Condensation Function of a <i>Z</i>,<i>Z</i>-Farnesyl Diphosphate Synthase.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure