Soman inhibition of butyrylcholinesterase in the presence of substrate: pressure and temperature perturbations.
Irreversible inhibition of butyrylcholinesterase by soman was studied in the presence of the substrate (o-nitrophenyl butyrate). Inhibition was found of the competitive complexing type. Study at different temperatures and pressures showed that the behavior of the enzyme differs from that of the inhibitor-free enzyme. In the absence of inhibitor, enzyme kinetics displayed a non-linear temperature dependence with a break at 21 degrees C. In the presence of a non-inhibitor structural analog of soman (pinacolyl dimethylphosphinate and methyl dimethylphosphinate), the Arrhenius plot break is slightly shifted (18 degrees C). On the other hand, in the presence of soman this break is abolished. The pressure-dependence of the substrate hydrolysis revealed also differences between the native enzyme and the enzyme in the presence of soman: the sign and magnitude of the apparent activation volume (delta V not equal to) were different for the two reactions. Beyond 300 bar, in the presence of soman, a plateau (delta V not equal to approx. 0) was observed over a large pressure range depending on temperature. Such a behavior with respect to temperature and pressure can reflect a soman-induced enzyme conformational state. Thus, temperature and pressure perturbations of the kinetics allow to complete the inhibition scheme of butyrylcholinesterase by soman. Our data suggest that upon soman binding, the enzyme undergoes a long-lived soman-induced-fit conformational change preceding the phosphonylation step. However, an alternative hypothesis according to which the enzyme processes a secondary soman-binding site cannot be ruled out.
Cléry
C
C
Centre de Recherches du Service de Santé des Armées, La Tronche, France.
Masson
P
P
Heiber-Langer
I
I
Balny
C
C
D001665:Binding Sites; D002091:Butyrylcholinesterase / Q000737:chemistry*; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D006868:Hydrolysis; D007700:Kinetics; D011312:Pressure; D011487:Protein Conformation; D012999:Soman / Q000494:pharmacology*; D013696:Temperature
D002800:Cholinesterase Inhibitors; D012999:Soman; D002091:Butyrylcholinesterase