Enhancement of Monascus yellow pigments production by activating the cAMP signalling pathway in Monascus purpureus HJ11.
Enhancement of Monascus yellow pigments production by activating the cAMP signalling pathway in Monascus purpureus HJ11.
复制标题
通过激活红曲红 HJ11 中的 cAMP 信号通路增强红曲黄色素的产生
DOI：
10.1186/s12934-020-01486-y
复制
发表时间：
2020-12-07
影响因子：
6.4
通讯作者：
Li M
中科院分区：
文献类型：
--
作者：
Liu J;Du Y;Ma H;Pei X;Li M
背景
由红曲霉产生的红曲霉氮杂环丙酮类色素(Monascus azapilone dyments，MonAZPs)，在日常生活中作为重要的食品着色剂和食品补充剂已被超过10亿人使用。此外，单偶氮化合物由于其多样的生理活性，近年来受到了更多的关注。然而，MonAzps的高微生物产量仍然不总是有保证的。在这里，本研究的目的是开发一种高效的单氮卓类生物工艺流程。
结果
在本研究中，外源环磷酸腺苷(CAMP)处理不仅诱导了MonAZPs的产生，而且还刺激了cAMP磷酸二酯酶基因mrPDE在紫云英HJ11中的表达。随后，用纯化的酶进行体外酶促反应，确定Mr PDE为cAMP磷酸二酯酶。此外，构建了mrPDE基因敲除突变体，以验证cAMP信号通路的激活。缺失mrPDE基因后，hj11细胞内cAMP浓度增加378%，PKA激酶活性增加1.5倍，表明cAMP信号通路被激活。ΔmrPDE菌株的单偶氮量为8563U/g，是WT菌株的2.3倍。此外，ΔmrPDE菌株的NAPDH/NADP+比值明显高于野生型菌株，这导致了黄色MonazP的比例较高。在补料分批发酵条件下，Δ的产量和产量分别达到332.1 U/m L和8739U/g。
结论
通过激活cAMP信号通路，成功地培育出一株高产Monazps的工程菌株。这项研究不仅描述了一种新的单氮唑类产生菌的开发策略，而且为在其他丝状真菌中生产次生代谢的工程努力提供了路线图。
Background
Monascus azaphilone pigments (MonAzPs), which were produced by Monascus species, have been used as important food colorant and food supplements for more than one billion people during their daily life. Moreover, MonAzPs recently have received more attention because of their diverse physiological activities. However, the high microbial production of MonAzPs is still not always guaranteed. Herein, the aim of this study was to develop an efficient biotechnological process for MonAzPs production.
Results
In this study, exogenous cyclic adenosine monophosphate (cAMP) treatment not only induced MonAzPs production, but also stimulated the expression of a cAMP phosphodiesterase gene, named as mrPDE, in M. purpureus HJ11. Subsequently, MrPDE was identified as a cAMP phosphodiesterase by in vitro enzymatic reaction with purified enzyme. Further, a gene knockout mutant of mrPDE was constructed to verify the activation of cAMP signalling pathway. Deletion of mrPDE in M. purpureus HJ11 improved cAMP concentration by 378% and enhanced PKA kinase activity 1.5-fold, indicating that activation of cAMP signalling pathway was achieved. The ΔmrPDE strain produced MonAzPs at 8563 U/g, with a 2.3-fold increase compared with the WT strain. Moreover, the NAPDH/NADP+ ratio of the ΔmrPDE strain was obviously higher than that of the wild type strain, which led to a higher proportion of yellow MonAzPs. With fed-batch fermentation of the ΔmrPDE strain, the production and yield of MonAzPs achieved 332.1 U/mL and 8739 U/g.
Conclusions
A engineered M. purpureus strain for high MonAzPs production was successfully developed by activating the cAMP signalling pathway. This study not only describes a novel strategy for development of MonAzPs-producing strain, but also provides a roadmap for engineering efforts towards the production of secondary metabolism in other filamentous fungi.
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