Kovarik_2004_Biochemistry_43_3222

Reference

Title : Mutant cholinesterases possessing enhanced capacity for reactivation of their phosphonylated conjugates - Kovarik_2004_Biochemistry_43_3222
Author(s) : Kovarik Z , Radic Z , Berman HA , Simeon-Rudolf V , Reiner E , Taylor P
Ref : Biochemistry , 43 :3222 , 2004
Abstract : Selective mutants of mouse acetylcholinesterase (AChE; EC 3.1.1.7) phosphonylated with chiral S(P)- and R(P)-cycloheptyl, -3,3-dimethylbutyl, and -isopropyl methylphosphonyl thiocholines were subjected to reactivation by the oximes HI-6 and 2-PAM and their reactivation kinetics compared with wild-type AChE and butyrylcholinesterase (EC 3.1.1.8). Mutations in the choline binding site (Y337A, Y337A/F338A) or combined with acyl pocket mutations (F295L/Y337A, F297I/Y337A, F295L/F297I/Y337A) were employed to enlarge active center gorge dimensions. HI-6 was more efficient than 2-PAM (up to 29000 times) as a reactivator of S(P)-phosphonates (k(r) ranged from 50 to 13000 min(-1) M(-1)), while R(P) conjugates were reactivated by both oximes at similar, but far slower, rates (k(r) < 10 min(-1) M(-1)). The Y337A substitution accelerated all reactivation rates over the wild-type AChE and enabled reactivation even of R(P)-cycloheptyl and R(P)-3,3-dimethylbutyl conjugates that when formed in wild-type AChE are resistant to reactivation. When combined with the F295L or F297I mutations in the acyl pocket, the Y337A mutation showed substantial enhancements of reactivation rates of the S(P) conjugates. The greatest enhancement of 120-fold was achieved with HI-6 for the F295L/Y337A phosphonylated with the most bulky alkoxy moiety, S(P)-cycloheptyl methylphosphonate. This significant enhancement is likely a direct consequence of simultaneously increasing the dimensions of both the choline binding site and the acyl pocket. The increase in dimensions allows for optimizing the angle of oxime attack in the spatially impacted gorge as suggested from molecular modeling. Rates of reactivation reach values sufficient for consideration of mixtures of a mutant enzyme and an oxime as a scavenging strategy in protection and treatment of organophosphate exposure.
ESTHER : Kovarik_2004_Biochemistry_43_3222
PubMedSearch : Kovarik_2004_Biochemistry_43_3222
PubMedID: 15023072

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Citations formats

Kovarik Z, Radic Z, Berman HA, Simeon-Rudolf V, Reiner E, Taylor P (2004)
Mutant cholinesterases possessing enhanced capacity for reactivation of their phosphonylated conjugates
Biochemistry 43 :3222

Kovarik Z, Radic Z, Berman HA, Simeon-Rudolf V, Reiner E, Taylor P (2004)
Biochemistry 43 :3222