葡萄糖-6-磷酸脱氢酶基因的Candida tropicalis过量表达及其对木糖醇合成代谢的影响

研究葡萄糖-6-磷酸脱氢酶基因g6pd过量表达对Candida tropicalis木糖醇生物合成代谢的影响。克隆Candidatropicalis CT16的g6pd基因，并将其与表达载体pYES-pgk重组连接，构建重组载体pYES-pgk-g6pd，LiAc/ssDNA/PEG方法转化导入C. tropicalis CT16，筛选阳性转化子，实现g6pd基因的过量表达。结果表明：发酵62 h，阳性转化子C. tropicalis SYG5的葡萄糖-6-磷酸脱氢酶活力提高了300%，发酵液中木糖醇质量浓度达到79.90 g/L，较野生型对照菌株的木糖醇产量提高了12.41%，木糖醇产率提高了44.94%。因此，葡萄糖-6-磷酸脱氢酶在C. tropicalis木糖醇的合成代谢途径中发挥重要作用，增强g6pd基因的表达，可以明显提高菌体NADPH供应量和还原力，有利于木糖醇的生物合成。

Cloning and Overexpression of Glucose-6-phosphate dehydrogenase in Candida tropicalis and Its Influence onXylitol Biosynthesis

The influence of overexpression of glucose 6-phosphate dehydrogenase (G6PDH) gene, g6pd, on xylitolbiosynthesis in Candida tropicalis was investigated. The gene g6pd was cloned from Candida tropicalis CT16 and insertedinto a yeast expression vector pYES-pgk, generating a recombinant expression vector pYES-pgk-g6pd, which was thenintroduced into C. tropicalis CT16 by the LiAc/ssDNA/PEG transformation method, resulting in over-expression of theg6pd gene. The fermentation results showed that the G6PDH activity was increased by 300%, and a maximum xylitol yieldof 79.90 g/L was achieved in the recombinant strain SYG5 harboring pYES-pgk-g6pd after 62 h of fermentation. Comparedto the wild type strain C. tropicalis CT16, the yield and productive rate of xylitol in strain SYG5 were increased by 11.30%and 44.91%, respectively. These results indicate that increasing the expression level of the gene g6pd significantly enhancesxylitol production and that the G6PDH plays a key role in the biosynthesis pathway of xylitol in C. tropicalis.