Vnitřní pH kvasinek a vliv transportu draselných iontů na jeho regulaci

• Main Author: Hošková, Barbora
• Other Authors: Urbánková, Eva
• Format: Dissertation
• Language: Czech
• Published: 2009
• Online Access: http://www.nusl.cz/ntk/nusl-278120

Cytosolic pH of yeast is regulated and maintained by cooperation of many transport proteins. A fluorescent protein pHluorin was used in this thesis to evaluate cytosolic pH of yeast Saccharomyces cerevisiae. pHluorin is sensitive to pH of its surrounding. The experiments were mostly carried out using spectrofluorometer, however also a method which enables using reader was improved and optimalized for future use. As well a specific arginin-phosphate growth medium was used. An influence of certain proteins on the ability to regulate cytosolic pH was also studied. Some proteins are connected with the regulation directly e. g. Pma1 ATPase, some of them in an indirect way e. g. Ypt6. It was proved, that the value of cytosolic pH depends on yeast strain and cultivation and experimental conditions. None of the gene deletions studied in this thesis changed regulation of pHcyt much, the cells were able to adjust pHcyt even in situation, when the external medium is acidified by sulphuric acid. Buffering capacity was measured with addition of weak base to the cell suspension. The values varied in range from 30 to 60 mM if the cells were cultivated in YNB growth medium. Cells cultivated in arginin-phosphate medium had higher buffering capacity.