| Autor: |
Ordentlich, A., Barak, D., Kronman, C., Benschop, H.P., Jong, L.P.A. de, Ariel, N., Barak, R., Segall, Y., Velan, B., Shafferman, A. |
| Jazyk: |
angličtina |
| Rok vydání: |
2000 |
| Předmět: |
|
| Zdroj: |
BioScience 2000, US Army Medical Defense Bioscience Review, 4-9 June 2000, 197 |
| Popis: |
Structural architecture of the human acetylcholinesterase (HuAChE) active center and the effect of adduct configurations on the aging process were explored through phosphylation and aging reaction of the AchE wild type and selected mutants using the four stereoisomers of 1,2,2-trimethylpropyl methylphosphonofluoridate (soman). Reactivity of wild type HuAChE toward the PS-soman diastereomers was 4.0-7.5x104-fold higher than toward the PR- diastereomers. Aging of the PSCS-somanyl-HuAChE conjugate was also >1.6x104-fold faster than that of the corresponding PRCS-somanyl adduct, as shown by both reactivation and electrospray mass spectrometry (ESI/MS) experiments. On the other hand, both processes exhibited very limited sensitivity to the chirality of the alkoxy group Cα carbon of either PS- or PR-diastereomers. Reactivities of HuAChE enzymes carrying replacements at the acyl pocket (F295A, F297A, F295L/F297V) indicate that stereoselectivity with respect to the soman phosphorus chirality depends on the structure of this binding subsite but this stereoselectivity cannot be explained only by limitation in the capacity to accommodate the PR- diastereomers. In addition, these acyl pocket enzyme mutants display some (5-10-fold) preference for the PRCR-soman over the PRCSstereomer, while reactivity of the hydrophobic pocket mutant enzyme W86F toward the PRCS-soman resembles that of the wild type HuAChE. Residue substitutions at the Hbond network (E202Q, E450A, Y133F, Y133A) and the hydrophobic pocket (F338A, W86A, W86F, Y337A) result in a limited stereoselectivity for the PSCS- over the PSCRstereomer. Aging of the PS-somanyl conjugates with all the HuAChE mutant enzymes tested, practically lacked stereoselectivity with respect to the Cα of the alkoxy moiety. Thus, the inherent asymmetry of the active center does not seem to affect the rate determining step of the dealkylation process, possibly because both the PSCS- and the PSCR-somanyl moieties yield the same carbocationic intermediate. This work was supported by the U.S. Army Research and Material Command under Contract DAMD17-96-C-6088. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
|
