Istituto di Biologia e Biotecnologia Agraria, Consiglio Nazionale delle Ricerche, 20133, Milano, Italy.
Istituto di Biofisica, Consiglio Nazionale delle Ricerche, 20133, Milano, Italy.
Physiol Plant. 2019 May;166(1):403-412. doi: 10.1111/ppl.12900. Epub 2019 Jan 20.
The cyanobacterium Synechocystis sp. PCC 6803 is a model species commonly employed for biotechnological applications. It is naturally able to accumulate zeaxanthin (Zea) and echinenone (Ech), but not astaxanthin (Asx), which is the highest value carotenoid produced by microalgae, with a wide range of applications in pharmaceutical, cosmetics, food and feed industries. With the aim of finding an alternative and sustainable biological source for the production of Asx and other valuable hydroxylated and ketolated intermediates, the carotenoid biosynthetic pathway of Synechocystis sp. PCC 6803 has been engineered by introducing the 4,4' β-carotene oxygenase (CrtW) and 3,3' β-carotene hydroxylase (CrtZ) genes from Brevundimonas sp. SD-212 under the control of a temperature-inducible promoter. The expression of exogenous CrtZ led to an increased accumulation of Zea at the expense of Ech, while the expression of exogenous CrtW promoted the production of non-endogenous canthaxanthin and an increase in the Ech content with a concomitant strong reduction of β-carotene (β-car). When both Brevundimonas sp. SD-212 genes were coexpressed, significant amounts of non-endogenous Asx were obtained accompanied by a strong decrease in β-car content. Asx accumulation was higher (approximately 50% of total carotenoids) when CrtZ was cloned upstream of CrtW, but still significant (approximately 30%) when the position of genes was inverted. Therefore, the engineered strains constitute a useful tool for investigating the ketocarotenoid biosynthetic pathway in cyanobacteria and an excellent starting point for further optimisation and industrial exploitation of these organisms for the production of added-value compounds.
集胞藻 PCC 6803 是一种常用于生物技术应用的模式生物。它天然能够积累玉米黄质（Zea）和岩藻黄质（Ech），但不能积累虾青素（Asx），虾青素是微藻产生的具有最高价值的类胡萝卜素，在制药、化妆品、食品和饲料行业有广泛的应用。为了寻找替代和可持续的生物来源来生产 Asx 和其他有价值的羟基化和酮化中间体，引入了 Brevundimonas sp. SD-212 的 4,4'β-胡萝卜素加氧酶（CrtW）和 3,3'β-胡萝卜素羟化酶（CrtZ）基因，这些基因受温度诱导启动子的控制，对集胞藻 PCC 6803 的类胡萝卜素生物合成途径进行了工程改造。外源性 CrtZ 的表达导致 Zea 的积累增加，而 Ech 的积累减少，而外源性 CrtW 的表达促进了非内源性的角黄素的产生，并增加了 Ech 的含量，同时强烈降低了 β-胡萝卜素（β-car）的含量。当共表达 Brevundimonas sp. SD-212 的两个基因时，获得了大量的非内源性 Asx，同时β-car 的含量也强烈下降。当 CrtZ 克隆在 CrtW 的上游时，Asx 的积累更高（约占总类胡萝卜素的 50%），但当基因位置颠倒时，仍然有显著的积累（约 30%）。因此，这些工程菌株为研究蓝细菌中酮类胡萝卜素生物合成途径提供了有用的工具，也是进一步优化和开发这些生物用于生产附加值化合物的良好起点。
