 ISSN 1392-0146 ISSN 2029-0578 (online) |
2010 m. Nr. 1-4 Expression of glucose dehydrogenase–synuclein hybrid in Saccharomyces cerevisiae
Yeast expression vectors bearing glucose dehydrogenase gene fused with α-Synuclein (GDHsyn) were constructed and successfully introduced into Saccharomyces cerevisiae cells. Optimization of growing conditions of yeast transformants and a partial purification on the CM column of hybrid proteins were performed. The effect of culturing conditions on glucose dehydrogenase activity of hybrid construct-bearing yeasts was determined. Evaluation of expression level and localization of GDHsynEco and GDHsynPvu proteins identified major GDH activity in the cytoplasmic fractions of both producents. The genetic construction of the hybrid and the growing conditions of producents was found to influence not only the level of protein production, but also its localization in the cell. GDHsynPvu and GDHsynEco hybrids are cleaved into individual components, and only traces of the hybrid protein are detectable on SDS-PAGE. Keywords: S. cerevisiae, synuclein, glucose dehydrogenase, hybrid protein |
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