A Novel Specific Inhibitor of Tetrapyrrole Biosynthesis in Plants
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The behavior of many proteins is known to be subject to allosteric regulation (changes in ligand binding or catalytic activity caused by binding of another molecule to a site on the protein that is different from the active site). This current invention is based on a novel model of allosteric regulation of protein function through modulation of an equilibrium of alternate, nonadditive, functionally distinct oligomeric assemblies (morpheein forms). The morpheein model of allostery, which requires oligomer dissociation, is illustrated using a morphing dice analogy. Different morpheein forms of a given protein have different surface characteristics that can be targeted for the development of a broad spectrum of bioactive agents.
This invention relates to a mechanism of inhibiting porphobilinogen synthase (PBGS). PBGS catalyzes a fundamental step in the biosynthesis of tetrapyrrole pigments, an activity that is essential to all organisms that carry out respiration, photosynthesis, or methanogenesis. PBGS has been shown to exist in an equilibrium of high-activity octamers and low-activity hexamers whose interconversion is at the level of two different dimer conformations.
In plants, PBGS resides in the chloroplast and is established to contain a physiologically relevant allosteric magnesium binding site. This allosteric site is present in the octameric form of the PBGS. Addition of magnesium draws the equilibrium to the octamer and increases the activity of the PBGS. Activation of PBGS through structural stabilization of the octamer may be counteracted through structural stabilization of the hexamer. A phylogenetically diverse surface cavity specific to hexamers is proposed as an inhibitor-binding site. Although both the magnesium-binding site on the octamer and the hexamer-specific small molecule binding site qualify as allosteric sites, their existence is mutually exclusive; the chemical characteristics of ligands that would bind at each of these two sites are disparate.
Small molecules selected in silico have been tested in vitro for their ability to stabilize the hexameric form of pea PBGS and to inhibit enzyme activity. Fox Chase Cancer Center investigators have discovered morphlock-1 – a potent inhibitor of pea PBGS that drives the PBGS oligomeric distribution dramatically toward the hexamer. Morphlock-1 does not inhibit the activity of human PBGS, nor does it alter the quaternary structure equilibrium of PBGS from humans, Drosophila melanogaster, Pseudomonas aeruginosa or Vibrio cholera. Thus, the demonstration that morphlock-1 is the specific inhibitor of pea (essential for photosynthesis), and not human PBGS (essential for heme biosynthesis), coupled with the phylogenetic variation in the inhibitor binding site, leads to the conclusion that PBGS is a viable target for the development of herbicides.
Lawrence, S.H., et al., "Shape lifting leads to small-molecule drug discovery," (2008) Chemistry & Biology 15:586-96
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