Identification of a Strong Quorum Sensing- and Thermo-Regulated Promoter for the Biosynthesis of a New Metabolite Pesticide Phenazine-1-carboxamide in Pseudomonas strain PA1201

Abstract

Phenazine-1-carboxamide (PCN) produced by multifarious Pseudomonas strains represents a promising candidate as a new metabolite pesticide due to its broad-spectrum antifungal activity and capacity to induce systemic resistance in plants. The rice rhizosphere Pseudomonas strain PA1201 contains two reiterated gene clusters, phz1 and phz2, for phenazine-1-carboxylic acid (PCA) biosynthesis; PCA is further converted into PCN by this strain using a functional phzH-encoding glutamine aminotransferase. However, PCN levels in PA1201 constitute approximately one-fifth of PCA levels and the optimal temperature for PCN synthesis is 28 °C. In this study, the phzH open reading frame (ORF) and promoter region were investigated and reannotated. phzH promoter P phzH was found to be a weak promoter, and PhzH levels were not sufficient to convert all of the native PCA into PCN. Following RNA Seq and promoter-lacZ fusion analyses, a strong quorum sensing (QS)-and thermo-regulated promoter P rhlI was identified and characterized. The activity of P phzH is approximately 1% of P rhlI in PA1201. After three rounds of promoter editing and swapping by P rhlI , a new PCNoverproducing strain UP46 was generated. The optimal fermentation temperature for PCN biosynthesis in UP46 was increased from 28 to 37 °C and the PCN fermentation titer increased 179.5-fold, reaching 14.1 g/L, the highest ever reported.

Activity of bacteria isolated from bats against Pseudogymnoascus destructans in China

“…Phenazine compounds can be extracted from various microorganisms and have been recognized as antibiotics against plant diseases, as well as inducing systemic resistance in plants (Santos Kron et al ., 2020 ; Jin et al ., 2020 ). PCA is a very important compound widely present in microbial metabolites from Pseudomonas , Streptomycetes and Bacillus spp.…”

H-NS family protein MvaU downregulates phenazine-1-carboxylic acid (PCA) biosynthesis via binding to an AT-rich region within the promoter of the phz2 gene cluster in the rhizobacterium Pseudomonas strain PA1201

“…Due to its broad-spectrum antibiotic activity and the ability to trigger plant immunity against pathogen attacks, PCA has been used as a major active component in the development of the new metabolite pesticide “Shenqinmycin” in China. As of 2016, 1% Shenqinmycin suspensions have been shown to prevent nine fungal plant diseases [ 40 ]. To improve PCA yields, the engineered strain PAIV was generated to achieve PCA yields of 9.8 g/L [ 25 ].…”

Synthetic strategies of phenazine derivatives: A review

“…Jin and co‐workers [129] developed an elegant formal synthesis of phenazine‐1‐carboxamide ( 264–265 ) using biosynthetic strategies. As shown in Scheme 29, using this methodology, firstly, they were identified and characterized a strong quorum sensing and thermo‐regulated promoter.…”

“…Reproduced with permission from Reference[128] (Copyright©2020 American Chemical Society) S C H E M E 2 9 Design of the biosynthetic pathway for compounds 264-265. Reproduced with permission from Reference[129] (Copyright©2020 American Chemical Society) conditions and short reaction times with excellent yields when compared to the established chemical oxidative methods.…”