Optimization of a heterologous pathway for the production of flavonoids from glucose
Optimization of a heterologous pathway for the production of flavonoids from glucose
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DOI：
10.1016/j.ymben.2011.02.002
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发表时间：
2011-07-01
影响因子：
8.4
通讯作者：
Stephanopoulos, Gregory
中科院分区：
文献类型：
--
作者：
Santos, Christine Nicole S.;Koffas, Mattheos;Stephanopoulos, Gregory
在过去的几年里，开发生产黄酮类化合物的高效微生物工艺一直是代谢工程的目标，主要是因为这些化合物据称具有促进健康的作用。尽管最近在提高菌种效价和产量方面取得了很大进展，但目前的发酵策略存在两大缺陷--(1)需要在培养基中添加昂贵的苯丙酸前体，(2)需要两种不同的培养基配方来生产生物量/蛋白质和类黄酮类化合物。在这项研究中，我们详细介绍了一系列能够绕过这两个问题的菌株的构建。将酪氨酸解氨酶(TAL)、4-香豆酸：辅酶A连接酶(4CL)、查尔酮合成酶(CHS)和查尔酮异构酶(CHI)四步异源途径组装到两个工程菌中，使其能够直接从葡萄糖中合成主要的类黄酮类前体柚皮苷元。在研究过程中，我们发现需要对酶的来源和相对基因表达水平进行广泛的优化，才能实现对香豆酸和柚皮苷的高质量积累。然而，一旦达到这种代谢平衡，这些菌株被发现能够从葡萄糖中产生29毫克/L的柚皮苷元，并在添加脂肪酸酶抑制剂蓝绿球蛋白的情况下，生产高达84毫克/L的柚皮苷元。这些结果是通过在单一的最小培养基配方中培养大肠杆菌而获得的，从而为开发一种简单且经济的直接从葡萄糖中生产类黄酮的微生物工艺铺平了道路。(C)2011 Elsevier Inc.保留所有权利。
The development of efficient microbial processes for the production of flavonoids has been a metabolic engineering goal for the past several years, primarily due to the purported health-promoting effects of these compounds. Although significant strides have been made recently in improving strain titers and yields, current fermentation strategies suffer from two major drawbacks-(1) the requirement for expensive phenylpropanoic precursors supplemented into the media and (2) the need for two separate media formulations for biomass/protein generation and flavonoid production. In this study, we detail the construction of a series of strains capable of bypassing both of these problems. A four-step heterologous pathway consisting of the enzymes tyrosine ammonia lyase (TAL), 4-coumarate:CoA ligase (4CL), chalcone synthase (CHS), and chalcone isomerase (CHI) was assembled within two engineered I.-tyrosine Escherichia coli overproducers in order to enable the production of the main flavonoid precursor naringenin directly from glucose. During the course of this investigation, we discovered that extensive optimization of both enzyme sources and relative gene expression levels was required to achieve high quantities of both p-coumaric acid and naringenin accumulation. Once this metabolic balance was achieved, however, such strains were found to be capable of producing 29 mg/l naringenin from glucose and up to 84 mg/l naringenin with the addition of the fatty acid enzyme inhibitor, cerulenin. These results were obtained through cultivation of E. coli in a single minimal medium formulation without additional precursor supplementation, thus paving the way for the development of a simple and economical process for the microbial production of flavonoids directly from glucose. (C) 2011 Elsevier Inc. All rights reserved.