Characterisation of geranylgeranyl diphosphate synthase from the sandfly Lutzomyia longipalpis

Ducker, Charles; French, Stanley; Pathak, Monika; Taylor, Harry; Sainter, Adam; Askem, William; Dreveny, Ingrid; Santana, Antônio Euzébio Goulart; Pickett, John A.; Oldham, Neil J.

doi:10.1016/j.ibmb.2023.104001

Cited by 3 publications

(4 citation statements)

References 52 publications

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“…Considering the essentiality of GGPPS, its inhibition by certain bisphosphonate molecules, and the fact of variability of residues involved in binding among insect species of interest, it could be hypothesized that lepidopteran GGPPS holds potential as a putative pesticide target. A GGPPS from Lutzomyia longipalpis GGPPS was also recently expressed and functionally characterised (Ducker et al, 2023). As GGPPS from humans and Plasmodium species have already been successfully used as a drug target (No et al, 2012;Zhang et al, 2013), close monitoring of species selectivity is required.…”

Section: Discussionmentioning

confidence: 99%

Characterisation of lepidopteran geranylgeranyl diphosphate synthase as a putative pesticide target

Katsavou,

Sarafoglou,

Balabanidou

et al. 2023

Insect Molecular Biology

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Geranylgeranyl pyrophosphate (diphosphate) synthase (GGPPS) plays an important role in various physiological processes in insects, such as isoprenoid biosynthesis and protein prenylation. Here, we functionally characterised the GGPPS from the major agricultural lepidopteran pests Spodoptera frugiperda and Helicoverpa armigera. Partial disruption of GGPPS by CRISPR in S. frugiperda decreased embryo hatching rate and larval survival, suggesting that this gene is essential. Functional expression in vitro of Helicoverpa armigera GGPPS in Escherichia coli revealed a catalytically active enzyme. Next, we developed and optimised an enzyme assay to screen for potential inhibitors, such as the zoledronate and the minodronate, which showed a dose‐dependent inhibition. Phylogenetic analysis of GGPPS across insects showed that GGPPS is highly conserved but also revealed several residues likely to be involved in substrate binding, which were substantially different in bee pollinator and human GGPPS. Considering the essentiality of GGPPS and its putative binding residue variability qualifies a GGPPS as a novel pesticide target. The developed assay may contribute to the identification of novel insecticide leads.

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Section: Discussionmentioning

confidence: 99%

Characterisation of lepidopteran geranylgeranyl diphosphate synthase as a putative pesticide target

Katsavou,

Sarafoglou,

Balabanidou

et al. 2023

Insect Molecular Biology

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“…Kinetic analysis was carried out using the EnzChek Pyrophosphate Assay Kit (Invitrogen E6645, Thermo Fisher) following a modified protocol for 96‐well plates (Barja & Rodríguez‐Concepci'on, 2020; Ducker et al, 2023). MhieGGPPS (5 μg) was mixed with different substrate combinations in buffer (25 mM Tris‐HCl, pH 7.4, 1 mM MgCl 2 and 50% glycerol, 200 μL total volume), then added 2‐amino‐6‐mercapto‐7‐methylpurine ribonucleoside (0.2 mM), purine nucleoside phosphorylase (0.2 U), andinorganic diphosphatase (0.006 U).…”

Section: Methodsmentioning

confidence: 99%

“…Three biological replicates were conducted for each group. In kinetic analyses, the V max and K m values of MhieGGPPS against IPP, DMAPP, ( E )‐GPP, and ( E )‐FPP were caculated using nonlinear regression model block in Michaelis–Menten equation model with GraphPad Prism 9.0 (Ducker et al, 2023). The V max and K m for IPP were determined in the range of 1–40 μM, with ( E )‐FPP concentration fixed at 40 μM.…”

Section: Methodsmentioning

confidence: 99%

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Functional characterization of a geranylgeranyl diphosphate synthase in the leaf beetle Monolepta hieroglyphica

Song,

Liu,

Dhiloo

et al. 2024

Arch Insect Biochem Physiol

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Geranylgeranyl diphosphate synthase (GGPPS) as the short‐chain prenyltransferases for catalyzing the formation of the acyclic precursor (E)‐GGPP has been extensively investigated in mammals, plants, and microbes, but its functional plasticity is poorly understood in insect species. Here, a single GGPPS in leaf beetle Monolepta hieroglyphica, MhieGGPPS, was functionally investigated. Phylogenetic analysis showed that MhieGGPPS was clustered in one clade with homologs and had six conserved motifs. Molecular docking results indicated that binding sites of dimethylallyl diphosphate (DMAPP), (E)‐geranyl pyrophosphate (GPP), and (E)‐farnesyl pyrophosphate (FPP) were in the chain‐length determination region of MhieGGPPS, respectively. In vitro, recombiant MhieGGPPS could catalyze the formation of (E)‐geranylgeraniol against different combinations of substrates including isopentenyl pyrophosphate (IPP)/DMAPP, IPP/(E)‐GPP, and IPP/(E)‐FPP, suggesting that MhieGGPPS could not only use (E)‐FPP but also (E)‐GPP and DMAPP as the allylic cosubstrates. In kinetic analysis, the (E)‐FPP was most tightly bound to MhieGGPPS than that of others. It was proposed that MhieGGPPS as a multifunctional enzyme is differentiated from the other GGPPSs in the animals and plants, which only accepted (E)‐FPP as the allylic cosubstrate. These findings provide valuable insights into understanding the functional plasticity of GGPPS in M. hieroglyphica and the novel biosynthesis mechanism in the isoprenoid pathway.

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A diterpene synthase from the sandfly Lutzomyia longipalpis produces the pheromone sobralene

Ducker,

Baines,

Guy

et al. 2024

Proc. Natl. Acad. Sci. U.S.A.

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The phlebotomine sandfly, Lutzomyia longipalpis , a major vector of the Leishmania parasite, uses terpene pheromones to attract conspecifics for mating. Examination of the L. longipalpis genome revealed a putative terpene synthase (TPS), which—upon heterologous expression in, and purification from, Escherichia coli —yielded a functional enzyme. The TPS, termed Ll TPS, converted geranyl diphosphate (GPP) into a mixture of monoterpenes with low efficiency, of which β-ocimene was the major product. ( E,E )-farnesyl diphosphate (FPP) principally produced small amounts of ( E )-β-farnesene, while ( Z,E )- and ( Z,Z )-FPP yielded a mixture of bisabolene isomers. None of these mono- and sesquiterpenes are known volatiles of L. longipalpis . Notably, however, when provided with ( E,E,E )-geranylgeranyl diphosphate (GGPP), Ll TPS gave sobralene as its major product. This diterpene pheromone is released by certain chemotypes of L. longipalpis , in particular those found in the Ceará state of Brazil. Minor diterpene components were also seen as products of the enzyme that matched those seen in a sandfly pheromone extract.

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Characterisation of geranylgeranyl diphosphate synthase from the sandfly Lutzomyia longipalpis

Cited by 3 publications

References 52 publications

Characterisation of lepidopteran geranylgeranyl diphosphate synthase as a putative pesticide target

Characterisation of lepidopteran geranylgeranyl diphosphate synthase as a putative pesticide target

Functional characterization of a geranylgeranyl diphosphate synthase in the leaf beetle Monolepta hieroglyphica

A diterpene synthase from the sandfly Lutzomyia longipalpis produces the pheromone sobralene

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