Metabolic Engineering of Synechocystis sp. PCC 6803 for Production of the Plant Diterpenoid Manoyl Oxide.
Metabolic Engineering of Synechocystis sp. PCC 6803 for Production of the Plant Diterpenoid Manoyl Oxide.
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DOI：
10.1021/acssynbio.5b00070
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发表时间：
2015-12-18
影响因子：
4.7
通讯作者：
Lindberg P
中科院分区：
文献类型：
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作者：
Englund E;Andersen-Ranberg J;Miao R;Hamberger B;Lindberg P
毛喉素是一种高价值的二萜类化合物，具有广泛的药物应用，天然存在于毛喉锦紫苏植物的根皮中。由于其复杂的分子结构，化学合成毛喉素在商业上没有吸引力。因此，从福斯克利氏植物中进行的劳动和资源密集型提取和纯化仍然是该化合物的当前来源。我们设计了单细胞蓝藻集胞藻属 sp。 PCC 6803 用于生产毛喉素前体 13R-马诺酰氧化物 (13R-MO)，为光驱动生物技术生产这种高价值化合物铺平了道路。在这项工作的过程中，开发了一系列用于集胞藻的新整合载体，并用于创建表达染色体整合的 CfTPS2 和 CfTPS3 的稳定系，CfTPS2 和 CfTPS3 是福斯克氏梭菌中负责形成 13R-MO 的酶。工程菌株产生的生产滴度高达 0.24 mg g–1 DCW 13R-MO。为了提高产量，通过引入来自毛喉棒状芽孢杆菌的选定酶，对 13R-MO 生产菌株进行了进一步改造，将滴度提高至 0.45 mg g–1 DCW。这项工作为进一步开发蓝细菌中复杂植物二萜类化合物的生产奠定了基础。
Forskolin is a high value diterpenoid with a broad range of pharmaceutical applications, naturally found in root bark of the plant Coleus forskohlii. Because of its complex molecular structure, chemical synthesis of forskolin is not commercially attractive. Hence, the labor and resource intensive extraction and purification from C. forskohlii plants remains the current source of the compound. We have engineered the unicellular cyanobacterium Synechocystis sp. PCC 6803 to produce the forskolin precursor 13R-manoyl oxide (13R-MO), paving the way for light driven biotechnological production of this high value compound. In the course of this work, a new series of integrative vectors for use in Synechocystis was developed and used to create stable lines expressing chromosomally integrated CfTPS2 and CfTPS3, the enzymes responsible for the formation of 13R-MO in C. forskohlii. The engineered strains yielded production titers of up to 0.24 mg g–1 DCW 13R-MO. To increase the yield, 13R-MO producing strains were further engineered by introduction of selected enzymes from C. forskohlii, improving the titer to 0.45 mg g–1 DCW. This work forms a basis for further development of production of complex plant diterpenoids in cyanobacteria.