Zamjena signalne sekvence i njezin utjecaj na aktivnost rekombinantnog proteina Pir2bla izloženog na površini stanične stijenke

undergraduate thesis

• Title (english): Replacement of signal sequence and its effect on the activity of Pir2bla recombinant protein exposed on the cell wall
• Author: Anita Duvnjak
• Mentor: Bojan Žunar (mentor)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Chemistry and Biochemistry); (Laboratory for Biochemistry); Zagreb
• Defense date and country: 2023-09-08, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Kvasac Saccharomyces cerevisiae može na svojoj staničnoj stijenci izložiti proteine, ukoliko posjeduju signalnu sekvencu koja ih usmjerava na površinu stanice. Međutim, slabo je istraženo kako zamjena izvorne signalne sekvence s drugom signalnom sekvencom utječe na efikasnost izlaganja proteina, kao i može li se takva strategija primijeniti za povećanu efikasnost izlaganja biotehnološki zanimljivih proteina na površinu stanične stijenke. U ovom radu istraženo je kako zamjena signalne sekvence nativnog proteina stanične stijenke Pir2 sa signalnom sekvencom proteina Ccw12 utječe na izlaganje fuzijskog proteina Pir2bla. U tu svrhu, nakon konstrukcije i uvođenja odgovarajućeg plazmida u soj kvasca BY 4741, kolorimetrijskim nitrocefinskim testom uspoređena je efikasnost izlaganja obiju inačica proteina. Rezultati testa pokazali su da kvasac S. cerevisiae na svojoj stijenci u prosjeku izlaže 17 % manje inačice proteina sa Ccw12 signalnom sekvencom, ali i da ova razlika u efikasnosti izlaganja nije statistički značajna (p = 0,15).

Abstract (english)

The yeast Saccharomyces cerevisiae can display on its cell wall proteins whose signalling sequence directs them to the cell's surface. However, little is known about how exchanging the original signalling sequence for another such sequence affects protein display, as well as whether this strategy could enhance the surface display of biotechnologically relevant proteins. In this study, we investigated how replacing the signalling sequence of the cell surface protein Pir2 with that of Ccw12 affects the surface display of the fusion protein Pir2bla. After constructing and introducing the appropriate plasmid into yeast strain BY 4741, the surface display efficiency of both protein versions was compared using a colorimetric nitrocefin test. The results showed that, on average, S. cerevisiae displayed 17% fewer proteins with the Ccw12 signalling sequence, although this difference in efficiency was not statistically significant (p = 0.15).

• Keywords: izlaganje heterolognih proteina; stanična stijenka; signalna sekvenca; Saccharomyces cerevisiae; β-laktamaza
• Keywords (english): surface display; cell wall; signaling sequence; Saccharomyces cerevisiae; β-lactamase
• Language: croatian
• URN:NBN: urn:nbn:hr:159:814123
• Study programme: Title: Biotechnology; Study programme type: university; Study level: undergraduate; Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2024-04-30 11:29:29