ABSTRACT
Applying the genomic library construction strategy and colony screening, a new aroA gene encoding 5-enolpyruvylshikimate-3-phosphate synthase has been identified, cloned and overexpressed in Escherichia coli, and the enzyme was purified to homogeneity. Kinetic analysis of the AroA( P.fluorescens ) indicated that the full-length enzyme exhibits 10-fold increased IC50 and an approximately 38-fold increased K ( i ) for glyphosate compared to those of the AroA( E.coli ), while retaining high affinity for the substrate phosphoenolpyruvate. Furthermore, we have transformed the new aroA ( P.fluorescens ) gene into Arabidopsis thaliana via a floral dip method, and demonstrated that transgenic A. thaliana plants exhibit significant glyphosate resistance when compared with the wild type.
Subject(s)
3-Phosphoshikimate 1-Carboxyvinyltransferase/genetics , Genes, Bacterial/genetics , Genomic Library , Pseudomonas fluorescens/enzymology , Pseudomonas fluorescens/genetics , 3-Phosphoshikimate 1-Carboxyvinyltransferase/chemistry , Adaptation, Physiological/drug effects , Amino Acid Sequence , Arabidopsis/genetics , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Genetic Vectors/genetics , Glycine/analogs & derivatives , Glycine/toxicity , Kinetics , Models, Molecular , Molecular Sequence Data , Phylogeny , Plant Roots/anatomy & histology , Plant Roots/drug effects , Plants, Genetically Modified , Pseudomonas fluorescens/drug effects , Pseudomonas fluorescens/growth & development , Sequence Alignment , Sequence Analysis, DNA , Transformation, Genetic/drug effects , GlyphosateABSTRACT
Trichlorophenol (TCP) and its derivatives are introduced into the environment through numerous sources, including wood preservatives and biocides. Environmental contamination by TCPs is associated with human health risks, necessitating the development of cost-effective remediation techniques. Efficient phytoremediation of TCP is potentially feasible because it contains a hydroxyl group and is suitable for direct phase II metabolism. In this study, we present a system for TCP phytoremediation based on sugar conjugation by overexpressing a Populus putative UDP-glc-dependent glycosyltransferase (UGT). The enzyme PtUGT72B1 displayed the highest TCP-conjugating activity among all reported UGTs. Transgenic Arabidopsis demonstrated significantly enhanced tolerances to 2,4,5-TCP and 2,4,6-TCP. Transgenic plants also exhibited a strikingly higher capacity to remove TCP from their media. This work indicates that Populus UGT overexpression in Arabidopsis may be an efficient method for phytoremoval and degradation of TCP. Our findings have the potential to provide a suitable remediation strategy for sites contaminated by TCP.
Subject(s)
Arabidopsis/genetics , Chlorophenols/metabolism , Populus/enzymology , Adaptation, Physiological/drug effects , Amino Acid Sequence , Arabidopsis/drug effects , Arabidopsis/physiology , Biodegradation, Environmental/drug effects , Chlorophenols/isolation & purification , Chlorophenols/toxicity , Chromatography, High Pressure Liquid , Genes, Bacterial/genetics , Glucosyltransferases/chemistry , Glucosyltransferases/genetics , Glucosyltransferases/metabolism , Glycosylation/drug effects , Humans , Kinetics , Models, Molecular , Molecular Sequence Data , Pichia/metabolism , Plants, Genetically Modified , Populus/genetics , Substrate Specificity/drug effectsABSTRACT
Phenols are toxic, environmentally persistent products of the chemical industry that are capable of bioaccumulation and biomagnifications in the food chain. Little is known of how plants respond to this compound. To understand the transcriptional changes under phenol, microarray experiments on Arabidopsis thaliana were performed. Microarray results revealed numerous perturbations in signaling and metabolic pathways. The results indicated that the phenol response was related to reactive oxygen species (ROS) accumulation and oxidative conditions, including ROS generated for pathogen defense.
