Jiang Lihong, Dong Chang, Liu Tengfei, Shi Yi, Wang Handing, Tao Zeng, Liang Yan, Lian Jiazhang
Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, China.
Hangzhou Global Scientific and Technological Innovation Center, Zhejiang University, Hangzhou, China.
Front Bioeng Biotechnol. 2021 Dec 9;9:764851. doi: 10.3389/fbioe.2021.764851. eCollection 2021.
Cytochrome P450 enzymes (P450s) are a superfamily of heme-thiolate proteins widely existing in various organisms and play a key role in the metabolic network and secondary metabolism. However, the low expression levels and activities have become the biggest challenge for P450s studies. To improve the functional expression of P450s in , an cDNA library was expressed in the betaxanthin-producing yeast strain, which functioned as a biosensor for high throughput screening. Three new target genes , , and were identified to improve the functional expression of in yeast, with accordingly the accumulation of betaxanthin increased for 1.32-, 1.86-, and 1.10-fold, respectively. In addition, these three targets worked synergistically/additively to improve the production of betaxanthin, representing a total of 2.36-fold improvement when compared with the parent strain. More importantly, these genes were also determined to effectively increase the activity of another P450 enzyme (CYP736A167), catalyzing the hydroxylation of -santalene to produce Z-α-santalol. Simultaneous overexpression of , , and increased -santalene to Z-α-santalol conversion rate for more than 2.97-fold. The present study reported a novel strategy to improve the functional expression of P450s in and promises the construction of platform yeast strains for the production of natural products.
细胞色素P450酶（P450s）是血红素硫醇盐蛋白的一个超家族，广泛存在于各种生物体中，在代谢网络和次生代谢中发挥关键作用。然而，低表达水平和活性已成为P450s研究的最大挑战。为了提高P450s在[具体生物]中的功能表达，一个cDNA文库在产虾青素的酵母菌株中表达，该酵母菌株作为高通量筛选的生物传感器。鉴定出三个新的靶基因[具体基因1]、[具体基因2]和[具体基因3]，以改善[具体P450s]在酵母中的功能表达，相应地，虾青素的积累分别增加了1.32倍、1.86倍和1.10倍。此外，这三个靶标协同/累加作用以提高虾青素的产量，与亲本菌株相比，总产量提高了2.36倍。更重要的是，还确定这些基因能有效提高另一种P450酶（CYP736A167）的活性，催化α-檀香烯的羟基化反应生成Z-α-檀香醇。同时过表达[具体基因1]、[具体基因2]和[具体基因3]可使α-檀香烯到Z-α-檀香醇的转化率提高超过2.97倍。本研究报道了一种提高P450s在[具体生物]中功能表达的新策略，并有望构建用于生产天然产物的平台酵母菌株。
