Naslov
Phosphorylation of mouse cholinesterases and mutants by DDVP and reactivation of conjugates by HI-6 and 2-PAM

Autori
Kovarik, Zrinka ; Wong, Lilly ; Radić, Zoran ; Taylor, Palmer

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Seventh International Meeting on Cholinesterases, Pucon, Čile / Inestrosa, Nibaldo C. - Pucón : P. Universidad Catolica de Chile, 2002, 64-65

Skup
Seventh International Meeting on Cholinesterases

Mjesto i datum
Pucón, Čile, 08.11.2002. - 12.11.2002

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
DDVP; mouse cholinesterase mutants; phosphorylation; reactivation

Sažetak
DDVP (O, O-dimethyl-O-(2, 2-dichlorovinyl) phosphate) inhibits cholinesterases yielding the smallest symmetrical phosphorylated enzyme conjugate. Five recombinant DNA-derived mouse acetylcholinesterases (AChE ; EC 3.1.1.7), containing the F295L, F297I and Y337A mutations, and mouse butyrylcholinesterase (BChE ; EC 3.1.1.8) were inhibited by DDVP. The phosphorylated enzymes have been analysed for reactivation by the oximes, HI-6 and 2-PAM. Both inhibition and reactivation rate constants were determined. Phosphorylation by DDVP was about 7-fold more rapid for BChE (ki = 4.5 x 105 min-1M-1) than for AChE (ki = 6.2 x 104 min-1M-1). Inhibition rates of mutants decreased slightly compared to wild type AChE with the largest reduction being 5-fold for F297I/Y337A. An exception was phosphorylation of F295L/Y337A mutant where the rate increased and approached the inhibition rate of BChE. The near complete reactivation (90-100%) with HI-6 and 2-PAM was achieved for all dimethoxyphosphoryl-cholinesterase conjugates. Both oximes reactivated BChE conjugate at a similar rate (kr(HI-6)=420 min-1M-1 ; kr(2-PAM)=360 min-1M-1). However, the BChE conjugate was reactivated about 4-fold faster by HI-6 and 1.5-fold faster by 2-PAM than the AChE conjugate. Partial substitution of AChE residues with residues found in BChE did not achieve an overall reactivation rate comparable to the phosphorylated BChE. The Y337A mutation, that enhanced k2, the maximal rate of reactivation, also increased Kox, the constant reflecting dissociation of the initial complex, thereby achieving no overall enhancement in rate. The double mutants were more resistant to oxime reactivation than the single mutants. In contrast to the phosphonates with more bulky alkoxy groups, enlarging the dimensions of the acyl pocket and choline binding site did not substantially enhance phosphorylation by DDVP or oxime mediated reactivation rates. (Supported by DAMD17-18014 grant to P.T.)

Izvorni jezik
Engleski

Znanstvena područja
Kemija

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