Characterisation of bacterial phosphonate transporters and their application as glyphosate biosensors

Phosphonates are compounds characterised by a direct C-P bond and are environmentally abundant, thought to comprise around 4% of the total phosphorus in some soils. This class of compounds contains abundant naturally occurring chemicals like 2-aminoethylphosphonate (2-AEP), and the important synthetic herbicide glyphosate. Bacteria have evolved systems to scavenge the phosphorus from these compounds to use as a nutrient. Transport of 2-AEP in most Gram-negative bacteria is mediated by an ABC transporter encoded by the phnCDE genes. The aim of this work was to discover and characterise phosphonate transporters present in bacteria, particularly ones that have been shown to uptake and catabolise glyphosate in the environment. A biochemical approach studying the substrate binding protein (SBP) component of ABC transporters revealed a number of proteins that can bind natural phosphonate with low µM affinity. Significantly, some of these appear also to recognise glyphosate and have been rationally engineered as potential scaffolds for a glyphosate biosensor. In parallel, a series of strains of the rhizosphere bacterium Sinorhizobium meliloti were tested for their ability to use different phosphonates as the sole phosphorus source. Deletion of the genes encoding the PhnCDE transporter limits the range of phosphonates that S. meliloti can grow on, but does not abolish growth on phosphonates altogether, and a further transporter deletion mutant revealed a 2-AEP specific ABC transporter in this organism. This work has expanded the knowledge of the transport of phosphonates in biology.