Alternate Pathways for Acetic Acid and Acetate Ion Release from Acetylcholinesterase

Abstract

Two competing passageways for the exit of acetic acid and acetate ion in Torpedo californica (Tc) acetylcholinesterase (AChE) were studied by examining free energies of passage through two potential trajectories using the umbrella sampling technique as implemented in CHARMM. The reaction coordinates were defined from Ser200 Oγ one through the 20 A long active site gorge ending with Trp279 and a 14 A long route ending at Arg244. The free energies were calculated in successive windows 0.5 A wide for 40–90 ps and the potential of mean force (PMF) was calculated along the reaction coordinate. The PMF for the migration of acetic acid decreases by § 8 kcal/mol after 9 A travel through the main gorge. The PMF profile for acetate ion migration falls to a 6 kcal/mol lower value than that of acetic acid in the main gorge. The free energy barrier for the migration of acetate ion is 1.5 kcal/mol due a constriction formed by Tyrl 21, Phe290, Phe330, Phe331 in the main gorge. The interaction between acetic acid/acetate ion and the OH group of Tyrl21 appears to guide their release through the main gorge. Acetate ion remains H-bonded to Tyrl21 until it approaches Trp279 when it is expelled into bulk water. Acetic acid encounters a 6 kcal/mol barrier through the alternate pathway, while the PMF for acetate ion drops ~ 27 kcal/mol when it approaches Arg244. Full MD simulations, free of restraint for 170 ps, result in the migration of acetate ion through the short channel but not through the main gorge. The results indicate that if acetic acid ionizes within 3.5 A from Ser200 Oγ, it would be more likely to exit via the alternate channel than through the main gorge.