Biyomühendislik Ana Bilim Dalı Tez Koleksiyonu

Permanent URI for this collectionhttps://hdl.handle.net/20.500.12573/417

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Browsing Biyomühendislik Ana Bilim Dalı Tez Koleksiyonu by Author "Çalış, Burak"

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    Recombinant production of GLP-1 analogue using escherichia coli host organism
    (Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2024) Çalış, Burak; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
    Diabetes is the most serious metabolic disorder correlated with obesity, hypertensionandcardiovascular conditions. High prevalence of Type II Diabetes Mellitus (T2DM) indicatesthe need for new medication development. In developing therapeutics, higher efficiencyand fewer adverse effect features are targeted primarily. Recombinant protein-basedbiotechnological drug molecules have been developed and used for the treatment of T2DM. Especially, GLP-1 analogues are known by their self-limiting mechanismandinsulinotropic effect. In this study, a novel GLP-1 analogue with increased stabilityandefficiency is produced using recombinant E. coli. The expression plasmid was constructedand confirmed by restriction digestion and whole plasmid sequencing. Then, itwastransformed into various E. coli strains followed by optimized lysis, growth and expressionconditions to maximize the yield of the GLP-1 analogue. Various parameters suchas preinduction time, induction point, induction IPTG concentration and post-inductiontemperature were tested for the succesfull expression with maximumyield. Consequently, it was achieved that E. coli BL21(DE3) as strain, 0.2 mM IPTG induction at OD600nmof 0.6and 18 °C overnight post-induction growth was the most promising conditions. Under theseconditions, the GLP-1 analogue was obtained in the insoluble fraction. Followingproteinanalysis and purification, quantification was performed and the highest titer of GLP-1analogue was measured as 626 µg/ml. As future prospect, using another host organismandchanging growth conditions can provide obtaining target protein in the soluble form.