The slow rate of inhibition of acetylcholinesterase by fluoride.

We measured the rate of reaction of fluoride with acetylcholinesterase using a stopped flow apparatus for measurements on the millisecond time scale with phenylacetate as a chromogenic substrate. We found that the second order rate constant is 5 X 10(3) liters/mol/sec, which is very slow for a small symmetric ion; it is 3-4 orders of magnitude smaller than for the substrates acetylcholine, acetylthiocholine, and phenylacetate. The slowness of this reaction suggests that fluoride does not find a preexisting binding site but must create one, probably by breaking and reforming hydrogen bonds. With hydrolysis measurements made on the usual time scale, we found kcat = 7.5 X 10(5) min-1 and KM = 2.0 mM. We also found that fluoride enhances substrate inhibition and that with low phenylacetate concentration the per cent inhibition is independent of substrate concentration.