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Acetylcholinesterases from the Disease Vectors Aedes aegypti and Anopheles gambiae: Functional Characterization and Comparisons with Vertebrate Orthologues
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
Vise andre og tillknytning
2015 (engelsk)Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, nr 10, artikkel-id e0138598Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Mosquitoes of the Anopheles (An.) and Aedes (Ae.) genus are principal vectors of human diseases including malaria, dengue and yellow fever. Insecticide-based vector control is an established and important way of preventing transmission of such infections. Currently used insecticides can efficiently control mosquito populations, but there are growing concerns about emerging resistance, off-target toxicity and their ability to alter ecosystems. A potential target for the development of insecticides with reduced off-target toxicity is the cholinergic enzyme acetylcholinesterase (AChE). Herein, we report cloning, baculoviral expression and functional characterization of the wild-type AChE genes (ace-1) from An. gambiae and Ae. aegypti, including a naturally occurring insecticide-resistant (G119S) mutant of An. gambiae. Using enzymatic digestion and liquid chromatography-tandem mass spectrometry we found that the secreted proteins were post-translationally modified. The Michaelis-Menten constants and turnover numbers of the mosquito enzymes were lower than those of the orthologous AChEs from Mus musculus and Homo sapiens. We also found that the G119S substitution reduced the turnover rate of substrates and the potency of selected covalent inhibitors. Furthermore, non-covalent inhibitors were less sensitive to the G119S substitution and differentiate the mosquito enzymes from corresponding vertebrate enzymes. Our findings indicate that it may be possible to develop selective non-covalent inhibitors that effectively target both the wild-type and insecticide resistant mutants of mosquito AChE.

sted, utgiver, år, opplag, sider
2015. Vol. 10, nr 10, artikkel-id e0138598
HSV kategori
Identifikatorer
URN: urn:nbn:se:umu:diva-110996DOI: 10.1371/journal.pone.0138598ISI: 000362511000010PubMedID: 26447952OAI: oai:DiVA.org:umu-110996DiVA, id: diva2:871890
Forskningsfinansiär
Swedish Research CouncilTilgjengelig fra: 2015-11-17 Laget: 2015-11-02 Sist oppdatert: 2018-06-07bibliografisk kontrollert
Inngår i avhandling
1. Towards Mosquitocides for Prevention of Vector-Borne Infectious Diseases: discovery and Development of Acetylcholinesterase 1 Inhibitors
Åpne denne publikasjonen i ny fane eller vindu >>Towards Mosquitocides for Prevention of Vector-Borne Infectious Diseases: discovery and Development of Acetylcholinesterase 1 Inhibitors
2016 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Alternativ tittel[sv]
Mot nya insekticider för bekämpning av sjukdomsbärande myggor : identifiering och utveckling av acetylkolinesteras 1 inhibitorer
Abstract [en]

Diseases such as malaria and dengue impose great economic burdens and are a serious threat to public health, with young children being among the worst affected. These diseases are transmitted by mosquitoes, also called disease vectors, which are able to transmit both parasitic and viral infections. One of the most important strategies in the battle against mosquito-borne diseases is vector control by insecticides and the goal is to prevent people from being bitten by mosquitoes. Today’s vector control methods are seriously threatened by the development and spread of insecticide-resistant mosquitos warranting the search for new insecticides. This thesis has investigated the possibilities of vector control using non-covalent inhibitors targeting acetylcholinesterase (AChE); an essential enzyme present in mosquitoes as well as in humans and other mammals. A key requirement for such compounds to be considered safe and suitable for development into new public health insecticides is selectivity towards the mosquito enzyme AChE1. The work presented here is focused on AChE1 from the disease transmitting mosquitoes Anopheles gambiae (AgAChE1) and Aedes aegypti (AaAChE1), and their human (hAChE) and mouse (mAChE) counterparts. By taking a medicinal chemistry approach and utilizing high throughput screening (HTS), new chemical starting points have been identified. Analysis of the combined results of three different HTS campaigns targeting AgAChE1, AaAChE1, and hAChE allowed the identification of several mosquito-selective inhibitors and a number of compound classes were selected for further development. These compounds are non-covalent inhibitors of AChE1 and thereby work via a different mechanism compared to current anti-cholinergic insecticides, whose activity is the result of a covalent modification of the enzyme. The potency and selectivity of two compound classes have been explored in depth using a combination of different tools including design, organic synthesis, biochemical assays, protein X-ray crystallography and homology modeling. Several potent inhibitors with promising selectivity for the mosquito enzymes have been identified and the insecticidal activity of one new compound has been confirmed by in vivo experiments on mosquitoes. The results presented here contribute to the field of public health insecticide discovery by demonstrating the potential of selectively targeting mosquito AChE1 using non-covalent inhibitors. Further, the presented compounds can be used as tools to study mechanisms important in insecticide development, such as exoskeleton penetration and other ADME processes in mosquitoes.

sted, utgiver, år, opplag, sider
Umeå: Umeå universitet, 2016. s. 123+27
Emneord
Mosquito, vector-borne diseases, vector control, insecticide, acetylcholinesterase, medicinal chemistry, high-throughput screening, organic synthesis, homology modeling, structure activity relationship, structure selectivity relationship
HSV kategori
Forskningsprogram
datorlingvistik; organisk kemi
Identifikatorer
urn:nbn:se:umu:diva-119924 (URN)978-91-7601-492-9 (ISBN)
Disputas
2016-05-27, KB3B1, KBC-huset, Umeå, 13:00 (engelsk)
Opponent
Veileder
Tilgjengelig fra: 2016-05-04 Laget: 2016-05-02 Sist oppdatert: 2018-06-07bibliografisk kontrollert
2. Selective inhibition of acetylcholinesterase 1 from disease-transmitting mosquitoes: design and development of new insecticides for vector control
Åpne denne publikasjonen i ny fane eller vindu >>Selective inhibition of acetylcholinesterase 1 from disease-transmitting mosquitoes: design and development of new insecticides for vector control
2017 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

Acetylcholinesterase (AChE) is an essential enzyme with an evolutionary conserved function: to terminate nerve signaling by rapid hydrolysis of the neurotransmitter acetylcholine. AChE is an important target for insecticides. Vector control by the use of insecticide-based interventions is today the main strategy for controlling mosquito-borne diseases that affect millions of people each year. However, the efficiency of many insecticides is challenged by resistant mosquito populations, lack of selectivity and off-target toxicity of currently used compounds. New selective and resistance-breaking insecticides are needed for an efficient vector control also in the future. In the work presented in this thesis, we have combined structural biology, biochemistry and medicinal chemistry to characterize mosquito AChEs and to develop selective and resistance-breaking inhibitors of this essential enzyme from two disease-transmitting mosquitoes.We have identified small but important structural and functional differences between AChE from mosquitoes and AChE from vertebrates. The significance of these differences was emphasized by a high throughput screening campaign, which made it evident that the evolutionary distant AChEs display significant differences in their molecular recognition. These findings were exploited in the design of new inhibitors. Rationally designed and developed thiourea- and phenoxyacetamide-based non-covalent inhibitors displayed high potency on both wild type and insecticide insensitive AChE from mosquitoes. The best inhibitors showed over 100-fold stronger inhibition of mosquito than human AChE, and proved insecticide potential as they killed both adult and larvae mosquitoes.We show that mosquito and human AChE have different molecular recognition and that non-covalent selective inhibition of AChE from mosquitoes is possible. We also demonstrate that inhibitors can combine selectivity with sub-micromolar potency for insecticide resistant AChE.

sted, utgiver, år, opplag, sider
Umeå: Umeå University, 2017. s. 67
Emneord
acetylcholinesterase, non-covalent inhibitor, vector control, insecticide, mosquito, vector-borne disease, high throughput screening, rational design
HSV kategori
Forskningsprogram
biokemi; datorlingvistik
Identifikatorer
urn:nbn:se:umu:diva-134625 (URN)978-91-7601-723-4 (ISBN)
Disputas
2017-06-02, KB.E3.03 (stora hörsalen), KBC-huset, Umeå, 09:00 (engelsk)
Opponent
Veileder
Tilgjengelig fra: 2017-05-12 Laget: 2017-05-10 Sist oppdatert: 2018-06-09bibliografisk kontrollert

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