Expression and subcellular localization of organophosphate hydrolase in acephate-degrading Pseudomonas sp. strain Ind01 and its use as a potential biocatalyst for elimination of organophosphate insecticides

Abstract

Significance and Impact of the Study: A biocatalyst capable of degrading a wide range of organophosphate (OP) insecticides was generated by expressing an organophosphate degradation gene in Pseudomonas sp. Ind01 involved in mineralization of acephate. The biocatalyst can be used to eliminate a wide range of OP insecticide residues from the environment. AbstractOrganophosphate hydrolase (OPH), the product of an organophosphatedegrading (opd) gene cloned from Brevundimonas diminuta, hydrolyses the triester linkage found in neurotoxic organophosphate (OP) insecticides and nerve agents. Despite the fact that OPHs have a broad substrate range, OP compounds with a P-S linkage, such as insecticides like acephate, are poor substrates for the enzyme. Expression of OPH in acephate-utilizing Pseudomonas sp. Ind01 generated a live biocatalyst capable of degrading a wide range of OP insecticides. The heterologously expressed OPH, which is a substrate of twin arginine transport (Tat) pathway, successfully targeted to the membrane of Pseudomonas sp. Ind01. The membrane-associated OPH had a size that coincided with the mature form of OPH (mOPH), suggesting successful processing and targeting of the expressed OPH to the membrane. Pseudomonas sp. Ind01 expressing OPH degraded a variety of OP insecticides besides using acephate as sole carbon source.

Antibiotic discovery throughout the Small World Initiative: A molecular strategy to identify biosynthetic gene clusters involved in antagonistic activity

“…Given the unique physical state of these distinct habitats, we reason that strains adapted to different environments should maintain unique metabolic pathways capable of producing diverse secondary metabolites that may exhibit antimicrobial effects against other bacteria. Moreover, pseudomonads have been shown to breakdown refractory recalcitrant compounds such as chloroanilines (Nitisakulkan et al., ), insecticides (Pinjari, Pandey, Kamireddy, & Siddavattam, ), and chitin (Thompson, Smith, Wilkinson, & Peek, ), inhibit the growth of pathogenic plant fungi (Nielsen, Sorensen, Fels, & Pedersen, ; Nielsen, Thrane, Christophersen, Anthoni, & Sorensen, ; Tran, Ficke, Asiimwe, Hofte, & Raaijmakers, ), exhibit antitumor activity (Ikeda et al., ; Ni et al., ), and inhibit growth of a wide range of bacteria including human pathogens of MRSA (Farrow & Pesci, ; Rode, Hanslo, de Wet, Millar, & Cywes, ), Mycobacterium tuberculosis (Gerard et al., ), collections of gram‐positive and gram‐negative bacteria (Ye et al., ), and P. aeruginosa isolated from cystic fibrosis patients (Chatterjee et al., ). Thus, Pseudomonas spp.…”

Environmental Pseudomonads Inhibit Cystic Fibrosis Patient-Derived Pseudomonas aeruginosa

“…Reflecting such genomic diversity is the collective capability to produce a widely diverse repertoire of secondary metabolites, including nonribosomal peptides, bacteriocins, and quinolones, that have been utilized for important roles ranging from bioremediation or biocontrol (see references 25 and 26 for a review) to the key precursor for the FDA-approved semisynthetic anticancer drug ET-743 (28). For example, pseudomonads have been shown to breakdown refractory recalcitrant compounds, such as chloroanilines (29), insecticides (30), and chitin (31), and they inhibit the growth of plant-pathogenic fungi (32–34), exhibit antitumor activity (35, 36), and inhibit the growth of a wide range of bacteria, including the human-pathogenic pathogens methicillin-resistant Staphylococcus aureus (37, 38) and Mycobacterium tuberculosis (39). Thus, unique selective pressures within soil and water environments likely contribute to the evolution of genomic diversity and the production of diverse natural metabolites that affect bacterial fitness and abundance in such habitats.…”

Degradation of Acephate and Its Intermediate Methamidophos: Mechanisms and Biochemical Pathways

“…Microbial degradation is a potential way to decontaminate pesticide-contaminated sites (Chen et al, 2015;Yang et al, 2018;Zhan et al, 2018;Huang et al, 2019;Bhatt et al, 2020a). Biodegradation microorganisms including bacteria, fungi, actinomycetes, yeasts, and algae can be obtained by enrichment culture and recombination technology (Pinjari et al, 2013;Chen et al, 2014;Birolli et al, 2019;Bhatt et al, 2020b). At present, many researchers are looking for effective acephate or methamidophos degrading microorganisms through enrichment culture, including sewage treatment systems, organophosphorus contaminated areas, industries, and agricultural fields (Chen et al, 2012;Gao et al, 2012;Li et al, 2014;Mohan and Naveena, 2015;Lin et al, 2016).…”

“…Through genetic engineering technology, the opd gene cloned from Brevundimonas mututa was isogenously expressed in the acephate-mineralizing strain Pseudomonas sp. Ind01, suggesting that the engineered strain could degrade a variety of OP insecticides (Pinjari et al, 2013).…”

“…Furthermore, there are many studies on the degradation pathways of acephate and methamidophos as well as studies on related functional genes and enzymes ( Li et al, 2007 ; Shen et al, 2010 ; Chino-Flores et al, 2012 ; Kumar et al, 2018 ). Microorganisms metabolize pesticides through their respective degradation pathways, and the metabolic capacity of microorganisms can be improved by recombination technology ( Pinjari et al, 2013 ; Arora et al, 2017 ; Cycoń et al, 2017 ; Zhang et al, 2018 ; Lin et al, 2020 ). Microbial degradation of acephate and methamidophos is a potential tool for large-scale pollutant removal.…”