On The Use Of The Experimentally Determined Enzyme Inhibition Constant As A Measure Of Absolute Binding Affinity

Abstract

Defined as a state function representing an inhibitors absolute affinity for its target enzyme, the experimentally determined enzyme inhibition constant (Ki) is widely used to rank order binding affinities of different inhibitors for a common enzyme or different enzymes for a common inhibitor and to benchmark computational approaches to predicting binding affinity. Herein, we report that adsorption of bis(7)-tacrine to the glass container surface increased its Ki against Electrophorus electricus acetylcholinesterase (eeAChE) to 3.2 {+/-} 0.1 nM (n = 5) compared to 2.9 {+/-} 0.4 pM (n = 5) that was determined using plastic containers with other assay conditions kept the same. We also report that, due to binding or \"adsorption\" of bis(7)-tacrine to the inactive eeAChE, the bis(7)-tacrine Ki increased from 2.9 {+/-} 0.4 pM (n = 5) to 734 {+/-} 70 pM (n = 5) as the specific eeAChE activity decreased from 342 U/mg to 26 U/mg while other assay conditions were kept the same. These results caution against using Kis to rank order binding potencies, define selectivity, or benchmark computational methods without knowing detailed assay conditions.\n\nAbbreviations