Development of Artificial Synthetic Pathway of Endophenazines in <i>Pseudomonas chlororaphis</i> P3

oleh: Ying Liu, Shengjie Yue, Muhammad Bilal, Malik Jan, Wei Wang, Hongbo Hu, Xuehong Zhang

Format: Article
Diterbitkan: MDPI AG 2022-02-01

Deskripsi

Endophenazine A is a terpenoid phenazine with phenazine-1-carboxylic acid (PCA), and dimethylallyl diphosphate (DMAPP) derived from the 2-methyl-D-erythritol-4-phosphate (MEP) pathway as the precursor, which shows good antimicrobial activity against several Gram-positive bacteria and fungi. However, the highest yield of endophenazine A was about 20 mg/L in <i>Streptomyces</i>, limiting its large-scale industrial development. <i>Pseudomonas chlororaphis</i> P3, possessing an efficient PCA synthesis and MEP pathways, is a suitable chassis to synthesize endophenazine A. Herein, we designed an artificial biosynthetic pathway for the synthesis of endophenazine A in <i>P. chlororaphis</i> P3. Primarily, the prenyltransferase PpzP from <i>Streptomyces anulatus</i> 9663 was introduced into <i>P. chlororaphis</i> P3 and successfully synthesized endophenazine A. Another phenazine compound, endophenazine A1, was discovered and identified as a leakage of the intermediate 4-hydroxy-3-methyl-2-butene pyrophosphate (HMBPP). Finally, the yield of endophenazine A reached 279.43 mg/L, and the yield of endophenazine A1 reached 189.2 mg/L by metabolic engineering and medium optimization. In conclusion, we successfully synthesized endophenazine A and endophenazine A1 in <i>P. chlororaphis</i> P3 for the first time and achieved the highest titer, which provides a reference for the heterologous synthesis of terpenoid phenazines.