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Revealing the hexokinase step of glycolysis in lactic acid producer fungus Rhizopus oryzae

2012-09-23
Alagoz, Eda
Acar, Seyda
Yucel, Meral
Hamamcı, Haluk
Rhizopus oryzae is a filamentous fungus which can grow on some renewable carbon sources and this makes it a good candidate for production of lactic acid and ethanol. However, in R. oryzae the production yields of these products are lower compared to alternative organisms. The utilization of the carbon source through glycolysis might end up in lactic acid or ethanol production. This makes the glycolytic pathway a focus of attention for our search for a way to increase the speed and efficiency of this pathway thus increasing the yield of desired end product. Hexokinase catalyse the first and irreversible step in glycolysis and implicated as one of the major control points in this pathway. In this study we are trying to investigate the regulation of glycolytic pathway of R. oryzae, in the light of studies done for Saccharomyces cerevisiae. The probable hexokinase genes were found from the annotated genome database of R. oryzae by doing a blast search using the known protein sequences of closely related S. cerevisiae hexokinases and glucokinase. Five of our ten probable hexokinase genes were obtained by using PCR. Sequencing results of three of them proved that these genes have no introns meaning they were expressed in R. oryzae. In the complementation study done in hexokinaless mutant of S. cerevisiae for one of these genes the growth of the transformant was observed on glucose or fructose containing media. Same experiments will be carried out for the rest of these genes.