The herbicidally active compound N-2-(5-chloro-pyridyl) aminomethylene bisphosphonic acid acts by inhibiting both glutamine and aromatic amino acid biosynthesis

The effect of the herbicidally active compound N-2-(5-chloro-pyridyl) amino methylene bisphosphonic acid (Cl-pyr-AMBPA), previously found in vitro to i nhibit the activity of the first enzyme in the shikimate pathway 3-deoxy-D- arabino-heptulosonate-7-phosphate synthase, was investigated in vivo on sus pension cultured cells of Nicotiana plumbaginifolia Viviani. Amino acid poo l measurement showed an actual reduction of tyrosine, tryptophan and phenyl alanine level following the addition of the compound to the growth medium. However, an even stronger effect was noticed for other amino acids, mainly glutamine. When the activity of the enzymes involved in the glutamate cycle was measured in the presence of Cl-pyr-AMBPA, glutamate synthase was unaff ected, while glutamine synthetase was significantly inhibited. Contrary to the herbicide phosphinothricin, the inhibitor bound reversibly to the enzym e. Kinetic analysis accounted for an inhibition of uncompetitive type with respect to ammonium, glutamate and ATP, with K-i values of 113, 97 and 39 m u M, respectively. Only the exogenous supply of a mixture of glutamine and aromatic amino acids relieved cell growth inhibition, suggesting that the p hytotoxic properties of Cl-pyr-AMBPA are due to inhibition of key enzymes i n both the corresponding pathways.