Eukaryotic OPA Anhydrolases

The ability of crude extracts of the protozoan Tetrahymena thermophila to hydrolyze the organophosphate DFP was discovered by Landis et al. (1985). Purification of the Tetrahymena material was conducted using a Sephacryl 

S-200 and S-300 molecular sizing column with a fraction volume of approximately half of that used in previous studies in order to increase resolution. Three repeatable peaks capable of the hydrolysis of DFP immediately became apparent. Upon the addition of Mn2+, a fourth peak appeared. The activities were identified as Tt DFPase-1, Tt DFPase-2. Tt DFPase-5, and their characteristics can be found in Table 10.2. Molecular weights of the Tetrahymena OPA anhydrases range from 67,000 Da to 96,000 Da. The activity of DFPase-4 is stimulated 17- to 30-fold with Mn2+. Tt DFPase-1, Tt DFPase-2, and Tt DFPase-3 are only stimulated two- to fourfold, and part of this increase may be due to contamination by the higher molecular weight Tt DFPase-4. Soman to DFP ratios are approximately 1 1 for the Tetrahymena OPA anhydrases. 

Mipafox is reversible and competitively inhibits Tt DFPase-1, Tt DFPase-2, and Tt DFPase-3 (Landis et al. 1989c). Hydrolysis of the mipafox by partially purified Tetrahymena extract was only 13% the rate of DFP. 

Mipafox is not inhibitory to the squid-type OPA anhydrase. As reported by Gay and Hoskin (1979), the active site prefers an isopropyl side chain compared to an ethyl or methyl group. 

Although the primary investigation into the OPA anhydrases of squid tissue has been of the squid-type OPA anhydrase, squid does contain the more widespread Mazur-type OPA anhydrase. Gill, heart, mantle, and blood tissues all exhibit OPA anhydrase activities that are Mn2+-stimulated and hydrolyze soman faster than DFP (Hoskin et al. 1984). 

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