3C86

OpdA from agrobacterium radiobacter with bound product diethyl thiophosphate from crystal soaking with tetraethyl dithiopyrophosphate- 1.8 A

Replaces:  2R1O

Summary for 3C86

• Entry DOI: 10.2210/pdb3c86/pdb
• Related: 2D2G 2D2H 2D2J 2R1K 2R1L 2R1M 2R1N 2R1P
• Descriptor: Phosphotriesterase, FE (II) ION, COBALT (II) ION, ... (6 entities in total)
• Functional Keywords: phosphotriesterase, opda, metalloenzyme, hydrolase
• Biological source: Agrobacterium tumefaciens
• Total number of polymer chains: 1
• Total formula weight: 35919.54

Authors

Ollis, D.L.,Jackson, C.J.,Foo, J.L.,Kim, H.K.,Carr, P.D.,Liu, J.W.,Salem, G. (deposition date: 2008-02-10, release date: 2008-02-19, Last modification date: 2023-11-15)

Primary citation

Jackson, C.J.,Foo, J.L.,Kim, H.K.,Carr, P.D.,Liu, J.W.,Salem, G.,Ollis, D.L.
In crystallo capture of a Michaelis complex and product-binding modes of a bacterial phosphotriesterase
J.Mol.Biol., 375:1189-1196, 2008

PubMed Abstract: 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.

PubMed: 18082180
DOI: 10.1016/j.jmb.2007.10.061

Experimental method

X-RAY DIFFRACTION (1.8 Å)