Naslov
Amino acid residues involved in the interaction of acetylcholinesterase and butyrylesterase with the carbamates Ro 02-0683 and bambuterol, and with terbutaline

Autori
Kovarik, Zrinka ; Radić, Zoran ; Grgas, Branka ; Škrinjarić-Špoljar, Mira ; Reiner, Elsa ; Simeon-Rudolf, Vera

Izvornik
Biochimica et biophysica acta. Protein structure and molecular enzymology (0167-4838) 1433 (1999), 1-2; 261-271

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
cholinesterases; inhibition; Ro 02-0683; bambuterol; terbutaline; modelling of enzyme/carbamate complexes
(Amino acid residues involved in the interaction of acetylcholinesterase and butyrylesterase with the carbamates Ro o2-0683 and bambuterol; and with terbutaline)

Sažetak
In order to identify amino acids involved in the interaction of acetylcholinesterase (AChE; EC 3.1.1.7) and butyrylcholinesterase (BChE; EC 3.1.1.8) with carbamates, the time course of inhibition of the recombinant mouse enzymes BChE w.t., AChE w.t. and of eleven site-directed AChE mutants by Ro 02-0683 and bambuterol was studied. In addition the reversible inhibition of cholinesterases by terbutaline, the leaving group of bambuterol, was studied. The bimolecular rate constant of AChE w.t. inhibition was 6.8 times smaller by Ro 02-0683 and 16,000 times smaller by bambuterol than that of BChE w.t.. The two carbamates were equipotent BChE inhibitors. Replacement of tyrosine 337 in AChE with alanine (resembling the choline binding site of BChE) resulted in 630 times faster inhibition by bambuterol. The same replacement decreased the inhibition by Ro 02-0683 ten times. The difference in size of the choline binding site in the two w.t. enzymes appeared critical for the selectivity of bambuterol and terbutaline binding. Removal of the charge with the mutation D74N caused a reduction in the reaction rate constants for Ro 02-0683 and bambuterol. Substitution of tyrosine 124 with glutamine in the AChE peripheral site significantly increased the inhibition rate for both carbamates. Substitution of phenylalanine 297 with alanine in the AChE acyl pocket decreased the inhibition rate by Ro 02-0683. Computational docking of carbamates provided plausible orientations of the inhibitors inside the active site gorge of mouse AChE and human BChE, thus substantiating involvement of amino acid residues in the enzyme active sites critical for the carbamate binding as derived from kinetic studies.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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