Sun Lei, Zeng Jia, Cui Peiwu, Wang Wei, Yu Dayu, Zhan Jixun
1Department of Biological Engineering, Utah State University, 4105 Old Main Hill, Logan, UT 84322-4105 USA.
2TCM and Ethnomedicine Innovation & Development Laboratory, School of Pharmacy, Hunan University of Chinese Medicine, Changsha, Hunan 410208 China.
J Biol Eng. 2018 Jun 7;12:9. doi: 10.1186/s13036-018-0103-x. eCollection 2018.
Regulatory genes play critical roles in natural product biosynthetic pathways. Chromomycins are promising anticancer natural products from actinomycetes. This study is aimed to create an efficient strain for production of these molecules by manipulating the regulatory genes.
A putative but silent chromomycin biosynthetic gene cluster was discovered in . Heterologous expression of the ketosynthase, chain length factor, and acyl carrier protein in confirmed that they are responsible for the assembly of a decaketide. Two regulatory genes are present in this gene cluster, including SARP-type activator SrcmRI and PadR-like repressor SrcmRII. Either overexpression of SrcmRI or disruption of SrcmRII turned on the biosynthetic pathway of chromomycins. The production titers of chromomycin A/A in R5 agar in these two strains reached 8.9 ± 1.2/13.2 ± 1.6 and 49.3 ± 4.3/53.3 ± 3.6 mg/L, respectively. An engineered strain was then constructed with both SrcmRII disruption and SrcmRI overexpression, which produced chromomycins A and A in R5 agar at 69.4 ± 7.6 and 81.7 ± 7.2 mg/L, respectively. Optimization of the culture conditions further increased the titers of chromomycins A and A respectively to 145.1 ± 15.3 and 158.3 ± 15.4 mg/L in liquid fermentation.
This work revealed the synergistic effect of manipulation of pathway repressor and activator genes in the engineering of a natural product biosynthetic pathway. The resulting engineered strain showed the highest production titers of chromomycins by a strain of , providing an efficient way to produce these pharmaceutically valuable molecules.
调控基因在天然产物生物合成途径中起关键作用。色霉素是来自放线菌的有前景的抗癌天然产物。本研究旨在通过操纵调控基因来创建一个高效生产这些分子的菌株。
在……中发现了一个推定的但沉默的色霉素生物合成基因簇。酮合成酶、链长因子和酰基载体蛋白在……中的异源表达证实它们负责十聚酮的组装。该基因簇中存在两个调控基因，包括SARP型激活剂SrcmRI和PadR样阻遏物SrcmRII。SrcmRI的过表达或SrcmRII的破坏均可开启色霉素的生物合成途径。这两种菌株在R5琼脂中色霉素A/A的产量分别达到8.9±1.2/13.2±1.6和49.3±4.3/53.3±3.6mg/L。然后构建了一个同时破坏SrcmRII和过表达SrcmRI的工程菌株，该菌株在R5琼脂中产生色霉素A和A的量分别为69.4±7.6和81.7±7.2mg/L。培养条件的优化进一步将液体发酵中色霉素A和A的产量分别提高到145.1±15.3和158.3±15.4mg/L。
这项工作揭示了在天然产物生物合成途径工程中操纵途径阻遏物和激活剂基因的协同作用。所得工程菌株在……菌株中显示出色霉素的最高产量，为生产这些具有药学价值的分子提供了一种有效方法。
