Enhanced biosynthesis of phenazine-1-carboxamide by engineered Pseudomonas chlororaphis HT66

Abstract

BackgroundPhenazine-1-carboxamide (PCN), a phenazine derivative, is strongly antagonistic to fungal phytopathogens. The high PCN biocontrol activity fascinated researcher’s attention in isolating and identifying novel bacterial strains combined with engineering strategies to target PCN as a lead molecule. The chemical route for phenazines biosynthesis employs toxic chemicals and display low productivities, require harsh reaction conditions, and generate toxic by-products. Phenazine biosynthesis using some natural phenazine-producers represent remarkable advantages of non-toxicity and possibly high yield in environmentally-friendlier settings.ResultsA biocontrol bacterium with antagonistic activity towards fungal plant pathogens, designated as strain HT66, was isolated from the rice rhizosphere. The strain HT66 was identified as Pseudomonas chlororaphis based on the colony morphology, gas chromatography of cellular fatty acids and 16S rDNA sequence analysis. The secondary metabolite produced by HT66 strain was purified and identified as PCN through mass spectrometry, and 1H, 13C nuclear magnetic resonance spectrum. The yield of PCN by wild-type strain HT66 was 424.87 mg/L at 24 h. The inactivation of psrA and rpeA increased PCN production by 1.66- and 3.06-fold, respectively, which suggests that psrA and rpeA are PCN biosynthesis repressors. qRT-PCR analysis showed that the expression of phzI, phzR, and phzE was markedly increased in the psrA and rpeA double mutant than in psrA or rpeA mutant. However, the transcription level of rpeA and rpeB in strain HT66ΔpsrA increased by 3.52- and 11.58-folds, respectively. The reduced psrA expression in HT66ΔrpeA strain evidenced a complex regulation mechanism for PCN production in HT66.ConclusionIn conclusion, the results evidence that P. chlororaphis HT66 could be modified as a potential cell factory for industrial-scale biosynthesis of PCN and other phenazine derivatives by metabolic engineering strategies.Electronic supplementary materialThe online version of this article (10.1186/s12934-018-0962-3) contains supplementary material, which is available to authorized users.

Genetic Engineering of Pseudomonas Chlororaphis Lzh-T5 to Enhance Production of Trans-2,3-Dihydro-3-Hydroxyanthranilic Acid

“…According to Chin-A-Woeng et al, PsrA played a negative regulatory role in the production of the antifungal metabolite PCA in P. chlororaphis PCL1391 [19]. Similar results were obtained from psrA disruption in P. chlororaphis HT66 [20]. According to our research, the sigma regulator psrA also exists in P. chlororaphis Lzh-T5.…”

“…RpeB was inhibition by RpeA, so an rpeA mutant strain enhanced the production of phenazine [23]. Similar results were observed from P. chlororaphis GP72 and P. chlororaphis HT66 [18,20]. The rpeA/rpeB TCST system was found in P. chlororaphis Lzh-T5, interruption of rpeA results in the increase in DHHA from 4.92 g/L to 5.52 g/L (Fig.…”

“…In P. chlororaphis HT66, GacA positively regulates the expression of psrA. psrA negatively controls the expression of rpoS and the expression of phenazine [20]. In this study, psrA was found in P. chlororaphis Lzh-T5.…”

“…While DHHA is an important intermediate product of phenazines, the production of phenazines is quite low. For example, in wild type P. chlororaphis GP72, the yield of 2-OH-PHZ is 4.5 mg/L, and the higher phenazine (PCN) production in wild type is about 400 mg/L in P. chlororaphis HT66 [20,21].…”

Generation of Xylose-inducible promoter tools forPseudomonasspecies and their use in implicating a role for the Type II secretion system protein XcpQ in inhibition of corneal epithelial wound closure

“…Pseudomonas species have been suggested for industrial production and bioremediation of molecules including, but not limited to, β-peptides (25), polyhdroxyalkanoates (26), phenazine-1-carboxyamide (27), proteases (28, 29), pseudofactin (30), rhamnolipids (31–33), silver nanoparticles (34), and toluene (35). The ability of P xut to activate expression of gene expression in the majority of cells and its relative strength in P. fluorescens compared to P BAD suggest that it may be useful for larger scale gene synthetic applications.…”