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Identifier 000443041
Title Protein biotechnology-based approaches for the functional and structural characterization of insect enzymes
Alternative Title Μέθοδοι πρωτεϊνικής βιοτεχνολογίας και στον λειτουργικό και δομικό χαρακτηρισμό πρωτεϊνών εντόμων
Author Σαράφογλου, Χαρά Δ.
Thesis advisor Βόντας, Ιωάννης
Μπαλαμπανίδου, Βασιλεία
Reviewer Κοκκινίδης, Μιχαήλ
Παυλίδης, Ιωάννης
Abstract Since the practice of agriculture first begun, the loss of yield due to several insect pests was apparent. Even in recent years, with advances in agricultural sciences, losses due to pests and diseases range from 10-90%. Not limited to agricultural damage, insects are also responsible for disease transmission having a profound impact on human and livestock health. To address economy and health issues caused by these insects, various field techniques are used for insect control that can be grouped into three categories: mechanical, biological, and chemical. Chemical methods are related to the use of insecticides targeting harmful insect species with the intention of eliminating them. Since the first applications of insecticides, insects, pressured by natural selection, started developing resistance mechanisms against them. This created the need of new methods of action in the form of new chemicals or new targets, forming this never-ending cycle. Protein biotechnology-based approaches is a powerful tool used to win the race against insecticide resistance. In vitro techniques are used to isolate and characterize existing or novel insecticide targets. Furthermore, resistance mechanisms can be studied in order to identify patterns and eventually resolve problems by slightly altering existing commercially available insecticides or even by designing new chemicals that by-pass insect defenses. For the purpose of this work, two such examples are being examined. As a new promising target, type III geranylgeranyl diphosphate synthase from Helicoverpa armigera was isolated and examined. After protein purification, an assay was adapted to measure activity and determine kinetic values. Through homology modeling, substrate binding patterns were evaluated, and new chemicals were designed using in silico techniques. These chemicals were later tested in the assay to determine potency by highthroughput screening. Finally, Cytochrome P450 from Bemicia tabaci, an enzyme with detoxification function, was studied as it is implicated in insecticide resistance. In this case, the main objective was to establish a reproducible protocol of bacterial expression and purification of the enzyme while keeping it functional in lab conditions. Due to this enzyme’s properties the above proved to be challenging and involved protein engineering strategies to improve efficiency.
Language English
Subject Geranylgeranyl diphosphate synthase
Insecticide targets
Pest control
Resistance mechanism
Αντιμετώπιση εντόμων
Μηχανισμοί ανθεκτικότητας
Μοριακή εντομολογία
Στόχοι εντομοκτόμων
Issue date 2021-11-26
Collection   School/Department--School of Sciences and Engineering--Department of Biology--Post-graduate theses
  Type of Work--Post-graduate theses
Permanent Link https://elocus.lib.uoc.gr//dlib/f/0/2/metadata-dlib-1634740404-298280-28316.tkl Bookmark and Share
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