Investigation of the binding of the Scw4 protein into the cell wall of yeast Saccharomyces cerevisiae

Abstract

Scw4 je protein koji se u staničnoj stijenci kvasca Saccharomyces cerevisiae veže kovalentno i nekovalentno, ali nije poznato koji dio gena kodira za regiju koja mu omogućuje kovalentno povezivanje. Provedeno je ispitivanje načina vezanja u stijenku nakon delecije dijelova gena SCW4 koji kodiraju za regije za koje se pretpostavlja da bi mogle omogućiti kovalentno povezivanje. U tu je svrhu kvasac uzgojen u selektivnoj YNB podlozi nakon čega su izolirane stijenke i njihovi proteini kuhanjem u SDS-u uz dodatak β-merkaptoetanola odnosno tretmanom stijenki s 30 mM NaOH. Izolirani proteini su zatim analizirani SDS elektroforezom po Laemmli-u i metodom Western blot. Prema dobivenim rezultatima može se pretpostaviti da se Scw4 protein u staničnu stijenku kvasca veže kovalentno preko regije koja je prema broju i redoslijedu glutaminskih ostataka slična repetitivnoj sekvenci PIR proteina. Dobivene rezultate je potrebno potvrditi daljnjim istraživanjima.Scw4 is a protein which can be bound covalently and non-covalently into the cell wall of yeast Saccharomyces cerevisiae, but it is not known which region of protein allows its covalent linkage. The way of its binding was examined after deletion of SCW4 gene segments encoding regions that could be responsible for covalent attachment of Scw4. For this purpose, the yeast was cultivated in a selective YNB medium and after that, the cell walls and proteins were isolated by boiling in SDS with β-mercaptoethanol or by treatment with a 30 mM NaOH. Isolated proteins were examined by SDS electrophoresis according to Laemmli and by Western blotting. According to the results, it can be assumed that the protein Scw4 is bound covalently via a region that has an arrangement and number of glutamine residues similar to that found in the repetitive sequence of PIR proteins that is responsible for its covalent binding. This results should be confirmed in further experiments