Репозиторий Университета

© 2018 Zlobin, Mokrushina, Terekhov, Zalevsky, Bobik, Stepanova, Aliseychik, Kartseva, Panteleev, Golovin, Belogurov, Gabibov and Smirnov. Butyrylcholinesterase (BChE) is considered as an efficient stoichiometric antidote against organophosphorus (OP) poisons. Recently we utilized combination of calculations and ultrahigh-throughput screening (uHTS) to select BChE variants capable of catalytic destruction of OP pesticide paraoxon. The purpose of this study was to elucidate the molecular mechanism underlying enzymatic hydrolysis of paraoxon by BChE variants using hybrid quantum mechanical/molecular mechanical (QM/MM) calculations. Detailed analysis of accomplished QM/MM runs revealed that histidine residues introduced into the acyl-binding loop are always located in close proximity with aspartate residue at position 70. Histidine residue acts as general base thus leading to attacking water molecule activation and subsequent SN2 inline hydrolysis resulting in BChE reactivation. This combination resembles canonical catalytic triad found in active centers of various proteases. Carboxyl group activates histidine residue by altering its pKa, which in turn promotes the activation of water molecule in terms of its nucleophilicity. Observed re-protonation of catalytic serine residue at position 198 from histidine residue at position 438 recovers initial configuration of the enzyme's active center, facilitating next catalytic cycle. We therefore suggest that utilization of uHTS platform in combination with deciphering of molecular mechanisms by QM/MM calculations may significantly improve our knowledge of enzyme function, propose new strategies for enzyme design and open new horizons in generation of catalytic bioscavengers against OP poisons.

© Bionika Media Ltd. Objective. To determine the expression level of the genes encoding the paraoxonase family enzymes (PON1, PON2, and PON3) in the placentas of women with uncomplicated pregnancy and in the development of preeclampsia Materials and methods. The investigation enrolled 26 pregnant women aged 24 to 35 years, including 14 women with normal pregnancy and 12 with preeclampsia. The expression of the PON1, PON2, and PON3 genes was analyzed by real-time PCR using the primers specific to these genes. Results. Women whose pregnancy was complicated by preeclampsia showed a significant decrease in the placental expression of the PON2 gene. The lowest PON2 gene expression was found in the placentas of women with severe preeclampsia. There were no significant differences in the placental expression level of the PON1 and PON3 genes in women with preeclampsia compared to healthy women. Conclusion. In women with preeclampsia, the placental PON2 gene expression decreases; however, the expression level of the PON1 and PON3 genes does not differ from that of the PON1 and PON3 genes in the placentas of patients with physiological pregnancy. The placental expression of the PON2 gene depends on the severity of preeclampsia; is lower in severe preeclampsia than in moderate preeclampsia.