Research of a specific competitive inhibitor of pyruvate dehydrogenase complex on Aurantiochytrium synthesizing fatty acids
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摘要: 本研究以合成乙酰辅酶A的丙酮酸脱氢酶复合体为扰动目标,通过应用底物特异竞争抑制剂——氟化丙酮酸(FP)来扰动乙酰辅酶A的合成,在考察扰动后裂殖壶菌的碳消耗、菌体生长、油脂合成、脂肪酸成分的基础上,分析了工业生产二十二碳六烯酸(DHA)菌株——裂殖壶菌中的乙酰辅酶A在普通脂肪酸合成途径(FAS途径)和类聚酮合酶途径(PKS途径)的分配比例变化。结果发现裂殖壶菌可转化利用FP;1 mmol/L FP在抑制总油脂合成的同时也显著改变了乙酰辅酶在FAS和PKS途径中的分配比率;降低了生物质合成,增强了二氧化碳生成量。表明裂殖壶菌的FAS途径对乙酰辅酶A的亲和力可能高于PKS途径;通过降低乙酰辅酶A供给量并不能提高总脂肪酸中多不饱和脂肪酸(DHA和DPA)的含量。这一结果解释了FAS途径和PKS途径的碳流分配调控机制,对于选育DHA高产菌株以及工业发酵过程优化具有重要意义。Abstract: Aurantiochytrium synthesizes polyunsaturated fatty acid( docosahexenoic acid( DHA) and docosapentaenoic acid( DPA)) through polyketide synthase( PKS) pathway,and saturated fatty acid( mainly palmic acid and oleic acid) through fatty acid synthase( FAS) pathway.Both pathways competitively used acetylCo A as carbon precursor. Understanding the mechanism to regulate the carbon partition between FAS pathway and PKS pathway was helpful for designation of high- DHA- yield strains and optimization of industrial fermentation process.In this research,the pyruvate dehydrogenase complex,which catalyzing the synthesis of acetyl- Co A,was treated by fluoropyruvate( FP)- a specific substrate- competitive inhibitor. Based on determination of carbon consumption,biomass,total oil accumulation and the distribution ratio of acetyl- Co A between FAS and PKS pathways were investigated.The results showed that FP could be assimilated and metabolized by Aurantiochytrium and FP could inhibit lipid synthesis and significantly change the partition ratioof acetyl- Co A between FAS and PKS pathways.Production of carbon- dioxide was enhanced by FP,but the accumulation of biomass was inhibited.These results hinted that the affinity of FAS pathway to acetyl- Co A was higher than that of PKS pathway and the content of polyunsaturated fatty acid( DHA and DPA) in total oil could not be increased by reducing supply of acetyl- Co A.
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Keywords:
- fluoropyruvate /
- docosahexenoic acid /
- pyruvate dehydrogenase complex /
- acyl-CoA /
- Aurantiochytrium
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