3LI4

Diisopropyl fluorophosphatase (DFPase), N120D,N175D,D229N mutant

Summary for 3LI4

• Entry DOI: 10.2210/pdb3li4/pdb
• Related: 3LI3 3LI5
• Descriptor: Diisopropyl-fluorophosphatase, CALCIUM ION (3 entities in total)
• Functional Keywords: beta propeller, calcium binding, phosphotriesterase, hydrolase
• Biological source: Loligo vulgaris (Common European squid)
• Total number of polymer chains: 1
• Total formula weight: 35201.85

Authors

Chen, J.C.-H. (deposition date: 2010-01-24, release date: 2010-04-07, Last modification date: 2023-09-06)

Primary citation

Blum, M.M.,Chen, J.C.
Structural characterization of the catalytic calcium-binding site in diisopropyl fluorophosphatase (DFPase)-Comparison with related beta-propeller enzymes.
Chem.Biol.Interact, 187:373-379, 2010

PubMed Abstract: The calcium-dependent phosphotriesterase diisopropyl fluorophosphatase (DFPase) from the squid Loligo vulgaris efficiently hydrolyzes a wide range of organophosphorus nerve agents. The two calcium ions within DFPase play essential roles for its function. The lower affinity calcium ion located at the bottom of the active site participates in the reaction mechanism, while the high affinity calcium in the center of the protein maintains structural integrity of the enzyme. The activity and structures of three DFPase variants targeting the catalytic calcium-binding site are reported (D121E, N120D/N175D/D229N, and E21Q/N120D/N175D/D229N), and the effect of these mutations on the overall structural dynamics of DFPase is examined using molecular dynamics simulations. While D229 is crucial for enzymatic activity, E21 is essential for calcium binding. Although at least two negatively charged side chains are required for calcium binding, the addition of a third charge significantly lowers the activity. Furthermore, the arrangement of these charges in the binding site is important for enzymatic activity. These results, together with earlier mutational, structural, and kinetic studies, show a highly evolved calcium-binding environment, with a specific electrostatic topology crucial for activity. A number of structural homologues of DFPase have been recently identified, including a chimeric variant of Paraoxonase 1 (PON1), drug resistance protein 35 (Drp35) from Staphylococcus aureus and the gluconolactonase XC5397 from Xanthomonas campestris. Surprisingly, despite low sequence identity, these proteins share remarkably similar calcium-binding environments to DFPase.

PubMed: 20206152
DOI: 10.1016/j.cbi.2010.02.043

Experimental method

X-RAY DIFFRACTION (1.35 Å)