Aminophosphonic Acid Derivatives 3r and 3s with Dual Herbicidal Activity and Crop Selectivity Targeted 5-Enolpyruvylshikimate-3-phosphate Synthase

In this study, a series of aminophosphoric acid derivatives were synthesized with 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) as a potential target, and their herbicidal activities were evaluated. At 600 g ai/ha, compounds 3r and 3s exhibited good herbicidal efficacy against Digitaria sanguinalis (DIGSA) and Setaria viridis (SETVI), with inhibition rates of 80% and 90%, and 80% and 95%, respectively. Additionally, both compounds 3r and 3s demonstrated significant enzyme inhibition activity against S. viridis EPSPS (SvEPSPS). Expression and purification of SvEPSPS and Portulaca oleracea EPSPS (PoEPSPS) proteins followed by microscale thermophoresis (MST) revealed that compounds 3r and 3s had excellent binding affinities for both SvEPSPS and PoEPSPS, with dissociation constants (K_d) of 0.65 and 1.27 μM, and 0.41 and 0.42 μM, respectively. Isothermal titration calorimetry (ITC) further validated the MST results, which were consistent with expectations. Molecular docking and molecular dynamics (MD) simulations further demonstrated that compounds 3r and 3s formed strong and stable complexes with SvEPSPS and PoEPSPS. Moreover, neither compound 3r nor 3s showed specific binding to the SvEPSPS^K355A and PoEPSPS^K355A mutant proteins, confirming that K355 is a key amino acid residue for binding. These findings suggest that compounds 3r and 3s have the potential to act as selective EPSPS inhibitors for the control of six species of grass weeds in crop fields.

5-enolpyruvylshikimate-3-phosphate synthase; aminophosphonic acid derivatives; crop selectivity; herbicidal activity; targeted protein.