Ispitivanje utjecaja prekomjerne ekspresije kompleksa GPI-transamidaze na ugradnju Ccw12bla proteina u staničnu stijenku kvasca S. cerevisiae

undergraduate thesis

• Title (english): Examination of the effect of overexpression of the GPI-transamidase complex on the incorporation of Ccw12bla protein into the cell wall of the yeast S. cerevisiae
• Author: Hana Pospiš
• Mentor: Renata Teparić (mentor)
• Mentor: Tea Martinić Cezar (pomoć pri izradi, neposredni voditelj)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Chemistry and Biochemistry); (Laboratory for Biochemistry); Zagreb
• Defense date and country: 2024-09-16, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Food Technology

Abstract

„Yeast surface display“ (YSD) sustav jednostavnija je, efikasnija i jeftinija metoda proizvodnje i imobilizacije enzima, nego klasične imobilizacije u kojima se za vezanje proteina na neki netopljivi nosač koriste kemikalije i postupci koje često imaju negativan utjecaj na enzimsku aktivnost. Rekombinantni protein mora sadržati protein od interesa fuzioniran sa nativnim proteinom stanične stijenke. Nedostatci ove metode su potencijalno smanjenje aktivnosti proteina i neuspješna ugradnja na površinu stanice. Bitna prepreka za industrijsku primjenu ove metode je njena još uvijek niska efikasnosti. Stoga se i dalje radi na tome da se efikasnost YSD poveća. Cilj ovog rada jest ispitati utjecaj prekomjerne ekspresije kompleksa GPI-transamidaze na ugradnju Ccw12bla proteina na površinu stanice. Ccw12bla protein eksprimiran je u stanicama kvasca divljeg tipa i u stanicama koje su bile transformirane plazmidima koji su sadržali genetičku informaciju za povećanu ekspresiju podjedinica kompleksa GPI-transamidaze. Uspješnost provedenog eksperimenta ispitana je mjerenjem aktivnosti β-laktamaze i imunoblotom ukupnih proteina stijenke.

Abstract (english)

“Yeast surface display" (YSD) system is a simpler, more efficient and cheaper method of enzyme production and immobilization than classic immobilizations in which chemicals and procedures used to attach proteins to some insoluble carrier often have a negative effect on enzyme activity. The recombinant protein must contain protein of interest fused to some native cell wall protein. The disadvantages of this method are the potential reduction of protein activity and unsuccessful incorporation on the cell surface. Low efficiency of YSD remains a significant obstacle for industrial application of this method, and efforts are still being made to improve its efficiency. The aim of this work is to examine the influence of overexpression of the GPI-transamidase complex on the incorporation of the Ccw12bla protein on the cell surface. The Ccw12bla protein was expressed in wild-type yeast cells and in cells transformed with plasmids containing genetic information for overexpression of the GPI transamidase complex subunits. The success of the conducted experiment was tested by measuring β-lactamase activity and immunoblot of cell wall proteins.

• Keywords: Ccw12bla protein; stanična stijenka; GPI-transamidaza; kvasac Saccharomyces cerevisiae; β-laktamaza
• Keywords (english): Ccw12bla protein; GPI-transamidase; cell wall; yeast Saccharomyces cerevisiae; β lactamase
• Language: croatian
• URN:NBN: urn:nbn:hr:159:573579
• Study programme: Title: Food Technology; Study programme type: university; Study level: undergraduate; Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka prehrambene tehnologije (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka prehrambene tehnologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2024-09-25 11:03:03