Characterization and genetic modification of Pseudomonas chlororaphis DZ15 with high yield of phenazine-1-carboxylic acid

Abstract Aims Phenazine-1-carboxylic acid (PCA) is an important secondary metabolite and a precursor for the biosynthesis of many phenazine derivatives, with highly efficient antifungal activity and environmentally friendly characteristics, which is mainly produced by microbial fermentation. Therefore, it is of great importance to isolate and identify new chassis strain with higher PCA production to meet the demand of PCA-efficient synthesis. In this study, a wild-type strain named DZ15 was initially characterized, which displayed the highest PCA production among all of wild-type strains reported. However, its physiological characteristics and metabolic performance remain unclear. Methods and results First, the taxonomic classification of strain DZ15 was initially identified through 16S rRNA-based phylogenetic analysis and subsequently classified as Pseudomonas chlororaphis subsp. aureofaciens. Then, genomic characteristics of P. chlororaphis DZ15 were investigated through comparative genomic analysis with P. chlororaphis GP72, LX24, and HT66 based on COG analysis and KEGG analysis. Furthermore, to explore the reasons for higher PCA production in P. chlororaphis DZ15, the expression level of the phenazine cluster promoter (Pphz) from P. chlororaphis DZ15 and other three strains (i.e. GP72, LX24, and HT66) was detected and compared in P. chlororaphis DZ15 and GP72 using a green fluorescent protein (GFP) fusion reporter. Among the four Pphz promoters, Pphz from P. chlororaphis DZ15 displayed the highest expression level, about 25413.44 AU/OD600. However, the expression level of all four promoters showed no significant difference in P. chlororaphis GP72 and much lower than their expression level in P. chlororaphis DZ15 with less than 5000 AU/OD600. Finally, the production of PCA was further improved from 370.14 mg·L−1 to 1532 mg·L−1 by deleting modifier gene phzO and negative regulatory genes rpeA, rsmE, and lon. Conclusion Our study firstly investigated the genomic characteristics of wild-type strain with the highest PCA production (i.e. P. chlororaphis DZ15) through whole-genome sequencing analysis. Then, our results indicated that the high expression level of phz cluster promoter and physiological characteristics of P. chlororaphis DZ15 both contributed to its high PCA production. Furthermore, PCA production in P. chlororaphis DZ15 was enhanced through genetic modification.