ACETYLCHOLINESTERASE - EFFECTS OF IONIC-STRENGTH AND DIMERIZATION ON THE RATE CONSTANTS

Brownian dynamics simulations are used to calculate diffusion-controll ed rate constants for the binding of a positively-charged ligand to ac etylcholinesterase (AChE) at 300 K, pH 7, and several ionic strengths. Models of the enzyme were constructed on the basis of the crystal str ucture of Torpedo californica AChE, and the ligand was modeled as a 5- angstrom sphere. Assignment of the charge distribution of the enzyme i s based on calculation of the fractional charges of its ionizable grou ps as a function of pH and ionic strength, by the finite difference Po isson-Boltzmann method. We find that the mean charge of the enzyme inc reases significantly with increasing ionic strength, with most of the increase occurring between 0 and 200 mM ionic strength. The charge dis tribution results in a very high dipole moment for the monomeric subun it of the protein: 1500 D relative to the Center of Diffusion. The mag nitude and orientation of the dipole moment are relatively insensitive to the ionic strength. At physiological ionic strength, electrostatic steering of the ligand increases the rate constant of the enzyme-liga nd encounter by more than one order of magnitude. The increase in prot ein charge with rising ionic strength weakens the ionic strength depen dence of the rate somewhat. The calculations reproduce the experimenta lly observed decrease of the rate constants with increasing ionic stre ngth. We observed no intrinisic rate difference for dimeric AChE as co mpared to the monomer: the rate constant of the dimer is twice that of the monomer.