| - | The mechanism by which the binuclear metallophosphotriesterases (PTEs, E.C. 3.1.8.1) catalyse substrate hydrolysis has been extensively studied., The mu-hydroxo bridge between the metal ions has been proposed to be the, initiating nucleophile in the hydrolytic reaction. In contrast, analysis, of some biomimetic systems has indicated that mu-hydroxo bridges are often, not themselves nucleophiles, but act as general bases for freely, exchangeable nucleophilic water molecules. Herein, we present, crystallographic analyses of a bacterial PTE from Agrobacterium, radiobacter, OpdA, capturing the enzyme-substrate complex during, hydrolysis. This model of the Michaelis complex suggests the alignment of, the substrate will favour attack from a solvent molecule terminally, coordinated to the alpha-metal ion. The bridging of both metal ions by the, product, without disruption of the mu-hydroxo bridge, is also consistent, with nucleophilic attack occurring from the terminal position. When, phosphodiesters are soaked into crystals of OpdA, they coordinate, bidentately to the beta-metal ion, displacing the mu-hydroxo bridge. Thus, alternative product-binding modes exist for the PTEs, and it is the, bridging mode that appears to result from phosphotriester hydrolysis., Kinetic analysis of the PTE and promiscuous phosphodiesterase activities, confirms that the presence of a mu-hydroxo bridge during phosphotriester, hydrolysis is correlated with a lower pK(a) for the nucleophile, consistent with a general base function during catalysis.
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| - | 2R1O is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Agrobacterium_tumefaciens Agrobacterium tumefaciens] with <scene name='pdbligand=FE2:'>FE2</scene>, <scene name='pdbligand=CO:'>CO</scene>, <scene name='pdbligand=DPJ:'>DPJ</scene> and <scene name='pdbligand=EDO:'>EDO</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Aryldialkylphosphatase Aryldialkylphosphatase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.1.8.1 3.1.8.1] Known structural/functional Sites: <scene name='pdbsite=AC1:Fe2+Binding+Site+For+Residue+A+800'>AC1</scene>, <scene name='pdbsite=AC2:Co+Binding+Site+For+Residue+A+801'>AC2</scene>, <scene name='pdbsite=AC3:Dpj+Binding+Site+For+Residue+A+1'>AC3</scene> and <scene name='pdbsite=AC4:Edo+Binding+Site+For+Residue+A+701'>AC4</scene>. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2R1O OCA].
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| - | In crystallo capture of a Michaelis complex and product-binding modes of a bacterial phosphotriesterase., Jackson CJ, Foo JL, Kim HK, Carr PD, Liu JW, Salem G, Ollis DL, J Mol Biol. 2008 Feb 1;375(5):1189-96. Epub 2007 Nov 1. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=18082180 18082180]
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