Doyle, Jason R., Burnell, James N., Haines, Dianne S., Llewellyn, Lyndon E., Motti, Cherie A., and Tapiolas, Dianne M. (2005) A rapid screening method to detect specific inhibitors of pyruvate orthophosphate dikinase as leads for C4 plant-selective herbicides. Journal of Biomolecular Screening, 10 (1). pp. 67-75.

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Abstract

Plants using the C4 photosynthetic pathway are highly represented among the world’s worst weeds, with only 4 C4 species being agriculturally productive (maize, sorghum, millet, and sugar cane). With the C4 acid cycle operating as a biochemical appendage of C3 photosynthesis, the additional enzymes involved in C4 photosynthesis represent an attractive target for the development of weed-specific herbicides. The rate-limiting enzyme of this metabolic pathway is pyruvate orthophosphate dikinase (PPDK). PPDK, coupled with phosphoenolpyruvate carboxylase and nicotinamide adenine dinucleotide-malate dehydrogenase, was used to develop a microplate-based assay to detect inhibitors of enzymes of the C4 acid cycle. The resulting assay had a Z' factor of 0.61, making it a high-quality assay able to reliably identify active test samples. Organic extracts of 6679 marine macroscopic organisms were tested within the assay, and 343 were identified that inhibited the 3 enzyme-coupled reaction. A high confirmation rate was achieved, with 95% of these hit extracts proving active again upon retesting. Sequential addition of phosphoenolpyruvate and oxaloacetate to the assay facilitated identification of 83 extracts that specifically inhibited PPDK.

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