title: A rapid screening method to detect specific inhibitors of pyruvate orthophosphate dikinase as leads for C4 plant-selective herbicides.
abstract: Plants using the C(4) photosynthetic pathway are highly represented among the world's worst weeds, with only 4 C(4) species being agriculturally productive (maize, sorghum, millet, and sugar cane). With the C(4) acid cycle operating as a biochemical appendage of C(3) photosynthesis, the additional enzymes involved in C(4) 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 C(4) 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.
authors.affiliation: Australian Institute of Marine Science, Townsville, Queensland, Australia.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Doyle
authors.forename: James N
authors.initials: JN
authors.lastname: Burnell
authors.forename: Dianne S
authors.initials: DS
authors.lastname: Haines
authors.forename: Lyndon E
authors.initials: LE
authors.lastname: Llewellyn
authors.forename: Cherie A
authors.initials: CA
authors.lastname: Motti
authors.forename: Dianne M
authors.initials: DM
authors.lastname: Tapiolas
mesh_terms: D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D006540:Herbicides / Q000737:chemistry / Q000494:pharmacology*; D008291:Malate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism; D015394:Molecular Structure; D019815:Oxalic Acid / Q000494:pharmacology; D010730:Phosphoenolpyruvate Carboxylase / Q000037:antagonists & inhibitors / Q000378:metabolism; D010936:Plant Extracts / Q000378:metabolism; D010944:Plants / Q000187:drug effects* / Q000201:enzymology*; D011772:Pyruvate, Orthophosphate Dikinase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013045:Species Specificity; D013997:Time Factors
chemical_list: D004791:Enzyme Inhibitors; D006540:Herbicides; D010936:Plant Extracts; D019815:Oxalic Acid; D008291:Malate Dehydrogenase; D011772:Pyruvate, Orthophosphate Dikinase; D010730:Phosphoenolpyruvate Carboxylase; D004121:Dimethyl Sulfoxide