体外苏氨酸循环固碳途径的构建  

In vitro Construction of the Threonine Cycle Pathway for Carbon Fixation

作　　者：巨晓芝 袁倩倩[2,3] 马春玲[2] 肖冬光[1] 马红武[2] 

机构地区：[1]天津科技大学生物工程学院,天津300457 [2]中国科学院系统微生物工程重点实验室中国科学院天津工业生物技术研究所,天津300308 [3]天津大学化工学院,天津300072

出　　处：《中国生物工程杂志》2017年第6期56-62,共7页China Biotechnology

基　　金：国家重点基础研究计划(2012CB725203);天津市科技支撑计划重点项目(14ZCZDSY00060);中科院重点部署项目(ZDRW-ZS-2016-3)资助项目

摘　　要：乙酰CoA是生物体代谢过程中重要的代谢物,也是许多有价值产品合成的前体物质。然而传统途径中通过丙酮酸脱羧生成乙酰CoA碳得率较低,因此构建一条高效的乙酰CoA合成途径具有重要的意义。由于在体外验证文献报道的高碳摩尔得率合成乙酰CoA的苏氨酸循环固碳途径,有较重要的理论意义和应用价值。因此在体外构建了苏氨酸循环固碳途径合成乙酰CoA,通过分段加酶的方式将其在体外进行了验证。在体外验证时,以丙酮酸为底物,则丝氨酸脱氨酶(Tdc B)为循环途径的最后一步反应。结果表明,当加入途径中除丝氨酸脱氨酶之外的酶时,测得的乙酰CoA浓度约1.5 mmol/L,待反应达到平衡时,加入丝氨酸脱氨酶,丝氨酸转化为丙酮酸,丙酮酸再次进入循环,乙酰CoA的量增加了约0.2 mmol/L,由此得出结论在体外苏氨酸循环实现了固碳。Acetyl-CoA is a central metabolite and an important precursor for the synthesis of many valuable products. However, the carbon yield of acetyl-CoA is relatively low due to the carbon loss at the pyruvate decarboxylation step. Therefore, it was of great significance to construct an efficient pathway for acetyl-CoA synthesis. The attempts is to validate the high carbon molar yield reported in the literature to synthesize acetyl- CoA threonine carbonization pathway, which has important theoretical significance and application value. So, carbon fixation pathway via threonine cycle was used to synthesize acetyl CoA in vitro. When pyruvate was used as the substrate, serine deaminase （TdcB） was the last step of the cycling pathway in the in vitro characterization. The results showed that when all enzymes except serine deaminase were added to the reaction system, the measured acetyl CoA concentration was about 1.5 mmol/L at an nearly equilibrium state. After serine deaminase was added, serine to pyruvate and re-entry into the circulation, acetyl CoA concentration was increased 0. 2 mmol/L. This result indicated that carbon fixation was achieved via the threonine cycle in vitro.

关 键 词：体外 固碳途径 苏氨酸循环 乙酰 COA 

分 类 号：Q815[生物学—生物工程]