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The (R)- and (S)- specific carbonyl reductases (RCR and SCR) with enhanced green fluorescence protein ( EGFP) were expressed in Saccharomyces cerevisiae W303-1A. By analysis of EGFP expression spectrum, the protein distribution and subcellular localization of two enzymes were determined.
By SOE-PCR method, the fused genes of RCR and SCR with EGFP were cloned and constructed on an eukaryotic expression vector pYX212, and transformed into S. cerevisiae by electroporation. With fluorescent protein as selection marker, the expression and distribution of RCR and SCR were observed.
The EGFP expression in S. cerevisiae cells was observed under the laser scanning confocal microscopy. It was showed that the two enzymes expressed stably in S. cerevisiae and mainly distributed in the intracellular membrane and cytoplasm, while minority were dotted in the center of cells. According to the fluorescent intensity, the expression level of SCR was much higher than that of RCR in S. cerevisiae. The biotransformation results sh